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1.
Nature ; 613(7945): 729-734, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36450357

RESUMO

Peptidoglycan and almost all surface glycopolymers in bacteria are built in the cytoplasm on the lipid carrier undecaprenyl phosphate (UndP)1-4. These UndP-linked precursors are transported across the membrane and polymerized or directly transferred to surface polymers, lipids or proteins. UndP is then flipped to regenerate the pool of cytoplasmic-facing UndP. The identity of the flippase that catalyses transport has remained unknown. Here, using the antibiotic amphomycin that targets UndP5-7, we identified two broadly conserved protein families that affect UndP recycling. One (UptA) is a member of the DedA superfamily8; the other (PopT) contains the domain DUF368. Genetic, cytological and syntenic analyses indicate that these proteins are UndP transporters. Notably, homologues from Gram-positive and Gram-negative bacteria promote UndP transport in Bacillus subtilis, indicating that recycling activity is broadly conserved among family members. Inhibitors of these flippases could potentiate the activity of antibiotics targeting the cell envelope.


Assuntos
Proteínas de Bactérias , Proteínas de Transporte , Sequência Conservada , Evolução Molecular , Bactérias Gram-Negativas , Bactérias Gram-Positivas , Fosfatos de Poli-Isoprenil , Antibacterianos/farmacologia , Bacillus subtilis/citologia , Bacillus subtilis/efeitos dos fármacos , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Bactérias Gram-Negativas/citologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/genética , Bactérias Gram-Negativas/metabolismo , Bactérias Gram-Positivas/citologia , Bactérias Gram-Positivas/efeitos dos fármacos , Bactérias Gram-Positivas/genética , Bactérias Gram-Positivas/metabolismo , Fosfatos de Poli-Isoprenil/metabolismo , Sintenia , Peptidoglicano/metabolismo , Parede Celular/química , Parede Celular/metabolismo
2.
Biomed Res Int ; 2020: 2562950, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32566673

RESUMO

The spermatozoa are transcriptionally dormant entities which have been recognized to be an archive of mRNA, coding for a variety of functionally crucial cellular proteins. This significant repository of mRNA is predicted to be associated with early embryogenesis and postfertilization. The mRNA transcripts which are tagged with minisatellites have been involved in the regulation of the gene functions as well as their organization. However, very little information is available regarding the expression of the transcripts tagged with minisatellites in spermatozoa. Therefore, in order to understand the functions and the conformational behavior of the proteins expressed from these minisatellite-tagged transcripts, we have performed a detailed in silico analysis using the sequences of the transcripts. The protein predicted from KF274549 showed the functionalities similar to uncharacterized C4orf26 proteins, while that obtained from KF274557 predicted to be a metallophosphoesterase. Furthermore, the structural folds in the structure of these predicted proteins were analyzed by using the homology modeling and their conformational behaviors in the explicit water conditions were analyzed by using the techniques of Molecular Dynamics (MD) simulations. This detailed analysis will facilitate the understanding of these proteins in the spermatozoon region and can be used for uncovering other attributes of the metabolic network.


Assuntos
Repetições Minissatélites/genética , Fosfoproteínas Fosfatases , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Humanos , Simulação de Dinâmica Molecular , Proteínas Nucleares/química , Proteínas Nucleares/classificação , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fosfoproteínas Fosfatases/química , Fosfoproteínas Fosfatases/classificação , Fosfoproteínas Fosfatases/genética , Fosfoproteínas Fosfatases/metabolismo , Conformação Proteica , Proteínas Proto-Oncogênicas c-akt
3.
Sci Rep ; 9(1): 12156, 2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31434971

RESUMO

The zebrafish sensory posterior lateral line is an excellent model system to study collective cell migration and organogenesis. Shootin1 is a cytoplasmic protein involved in neuronal polarization and axon guidance. Previous studies have shown that shootin1 couples actin filament retrograde flow with extracellular adhesive substrates at the leading edge of axonal growth cones, thereby producing mechanical force for the migration and guidance of axonal growth cones. However, the functions of shootin in peripheral cells remain unknown. Here we identified two novel shootin family members, shootin2 and shootin3. In zebrafish, shootin1 and shootin3 are expressed in the posterior lateral line primordium (PLLP) and neuromasts during embryonic development. A shootin1 mutant displayed a reduced speed of PLLP migration, while shootin1;shootin3 double mutation inhibited cell proliferation in the PLLP. Furthermore, our results suggest that shootin1 and shootin3 positively regulate the number of neuromasts and the number of cells in deposited neuromasts. Our study demonstrates that shootins mediate collective cell migration of the posterior lateral line primordium and formation of neuromasts in zebrafish.


Assuntos
Proteínas de Transporte/metabolismo , Sistema da Linha Lateral/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/crescimento & desenvolvimento , Actinas/metabolismo , Animais , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Movimento Celular , Desenvolvimento Embrionário , Edição de Genes , Microscopia de Fluorescência , Neurônios/fisiologia , Organogênese , Filogenia , Ligação Proteica , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/classificação , Proteínas de Peixe-Zebra/genética
4.
J Biol Chem ; 294(37): 13545-13559, 2019 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-31341017

RESUMO

The homeodomain-interacting protein kinase (HIPK) family is comprised of four nuclear protein kinases, HIPK1-4. HIPK proteins phosphorylate a diverse range of transcription factors involved in cell proliferation, differentiation, and apoptosis. HIPK2, thus far the best-characterized member of this largely understudied family of protein kinases, plays a role in the activation of p53 in response to DNA damage. Despite this tumor-suppressor function, HIPK2 is also found overexpressed in several cancers, and its hyperactivation causes chronic fibrosis. There are currently no structures of HIPK2 or of any other HIPK kinase. Here, we report the crystal structure of HIPK2's kinase domain bound to CX-4945, a casein kinase 2α (CK2α) inhibitor currently in clinical trials against several cancers. The structure, determined at 2.2 Å resolution, revealed that CX-4945 engages the HIPK2 active site in a hybrid binding mode between that seen in structures of CK2α and Pim1 kinases. The HIPK2 kinase domain crystallized in the active conformation, which was stabilized by phosphorylation of the activation loop. We noted that the overall kinase domain fold of HIPK2 closely resembles that of evolutionarily related dual-specificity tyrosine-regulated kinases (DYRKs). Most significant structural differences between HIPK2 and DYRKs included an absence of the regulatory N-terminal domain and a unique conformation of the CMGC-insert region and of a newly defined insert segment in the αC-ß4 loop. This first crystal structure of HIPK2 paves the way for characterizing the understudied members of the HIPK family and for developing HIPK2-directed therapies for managing cancer and fibrosis.


Assuntos
Proteínas de Transporte/química , Proteínas Serina-Treonina Quinases/química , Sequência de Aminoácidos , Sítios de Ligação , Proteínas de Transporte/classificação , Proteínas de Transporte/metabolismo , Domínio Catalítico , Cristalografia por Raios X , Evolução Molecular , Humanos , Simulação de Dinâmica Molecular , Naftiridinas/química , Naftiridinas/metabolismo , Fenazinas , Filogenia , Ligação Proteica , Proteínas Serina-Treonina Quinases/classificação , Proteínas Serina-Treonina Quinases/metabolismo , Estrutura Terciária de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Alinhamento de Sequência
5.
J Proteome Res ; 18(5): 2139-2159, 2019 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-30985132

RESUMO

Macrophages are involved in the primary human response to Candida albicans. After pathogen recognition, signaling pathways are activated, leading to the production of cytokines, chemokines, and antimicrobial peptides. ATP binding proteins are crucial for this regulation. Here, a quantitative proteomic and phosphoproteomic approach was carried out for the study of human macrophage ATP-binding proteins after interaction with C. albicans. From a total of 547 nonredundant quantified proteins, 137 were ATP binding proteins and 59 were detected as differentially abundant. From the differentially abundant ATP-binding proteins, 6 were kinases (MAP2K2, SYK, STK3, MAP3K2, NDKA, and SRPK1), most of them involved in signaling pathways. Furthermore, 85 phosphopeptides were quantified. Macrophage proteomic alterations including an increase of protein synthesis with a consistent decrease in proteolysis were observed. Besides, macrophages showed changes in proteins of endosomal trafficking together with mitochondrial proteins, including some involved in the response to oxidative stress. Regarding cell death mechanisms, an increase of antiapoptotic over pro-apoptotic signals is suggested. Furthermore, a high pro-inflammatory response was detected, together with no upregulation of key mi-RNAs involved in the negative feedback of this response. These findings illustrate a strategy to deepen the knowledge of the complex interactions between the host and the clinically important pathogen C. albicans.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Candida albicans/crescimento & desenvolvimento , Proteínas de Transporte/genética , Interações Hospedeiro-Patógeno , Proteínas Mitocondriais/genética , Fosfoproteínas/genética , Trifosfato de Adenosina/imunologia , Trifosfato de Adenosina/metabolismo , Proteínas Reguladoras de Apoptose/classificação , Proteínas Reguladoras de Apoptose/imunologia , Candida albicans/patogenicidade , Proteínas de Transporte/classificação , Proteínas de Transporte/imunologia , Morte Celular/genética , Morte Celular/imunologia , Retroalimentação Fisiológica , Humanos , Marcação por Isótopo , Proteínas Mitocondriais/classificação , Proteínas Mitocondriais/imunologia , Fagocitose/imunologia , Fosfopeptídeos/química , Fosfopeptídeos/isolamento & purificação , Fosfoproteínas/classificação , Fosfoproteínas/imunologia , Biossíntese de Proteínas , Mapeamento de Interação de Proteínas , Proteômica/métodos , Transdução de Sinais , Células THP-1
6.
Cell ; 174(1): 156-171.e16, 2018 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-29909984

RESUMO

Extracellular proTGF-ß is covalently linked to "milieu" molecules in the matrix or on cell surfaces and is latent until TGF-ß is released by integrins. Here, we show that LRRC33 on the surface of microglia functions as a milieu molecule and enables highly localized, integrin-αVß8-dependent TGF-ß activation. Lrrc33-/- mice lack CNS vascular abnormalities associated with deficiency in TGF-ß-activating integrins but have microglia with a reactive phenotype and after 2 months develop ascending paraparesis with loss of myelinated axons and death by 5 months. Whole bone marrow transplantation results in selective repopulation of Lrrc33-/- brains with WT microglia and halts disease progression. The phenotypes of WT and Lrrc33-/- microglia in the same brain suggest that there is little spreading of TGF-ß activated from one microglial cell to neighboring microglia. Our results suggest that interactions between integrin-bearing cells and cells bearing milieu molecule-associated TGF-ß provide localized and selective activation of TGF-ß.


Assuntos
Proteínas de Transporte/metabolismo , Microglia/metabolismo , Sistema Nervoso/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Axônios/metabolismo , Transplante de Medula Óssea , Encéfalo/metabolismo , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Células Cultivadas , Integrinas/metabolismo , Estimativa de Kaplan-Meier , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/citologia , Mutagênese Sítio-Dirigida , Doenças Neurodegenerativas/mortalidade , Doenças Neurodegenerativas/patologia , Doenças Neurodegenerativas/terapia , Filogenia , Ligação Proteica , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Fator de Crescimento Transformador beta/genética
7.
Biochim Biophys Acta Bioenerg ; 1858(8): 641-654, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28342810

RESUMO

Cancer cells undergo major changes in energy and biosynthetic metabolism. One of them is the Warburg effect, in which pyruvate is used for fermentation rather for oxidative phosphorylation. Another major one is their increased reliance on glutamine, which helps to replenish the pool of Krebs cycle metabolites used for other purposes, such as amino acid or lipid biosynthesis. Mitochondria are central to these alterations, as the biochemical pathways linking these processes run through these organelles. Two membranes, an outer and inner membrane, surround mitochondria, the latter being impermeable to most organic compounds. Therefore, a large number of transport proteins are needed to link the biochemical pathways of the cytosol and mitochondrial matrix. Since the transport steps are relatively slow, it is expected that many of these transport steps are altered when cells become cancerous. In this review, changes in expression and regulation of these transport proteins are discussed as well as the role of the transported substrates. This article is part of a Special Issue entitled Mitochondria in Cancer, edited by Giuseppe Gasparre, Rodrigue Rossignol and Pierre Sonveaux.


Assuntos
Proteínas de Transporte/fisiologia , Mitocôndrias/metabolismo , Proteínas Mitocondriais/fisiologia , Neoplasias/metabolismo , Transporte Biológico , Proteínas de Transporte/biossíntese , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Metabolismo Energético/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas Mitocondriais/biossíntese , Proteínas Mitocondriais/classificação , Proteínas Mitocondriais/genética , Modelos Biológicos , Família Multigênica , Neoplasias/genética , Neoplasias/patologia , Projetos de Pesquisa , Células Tumorais Cultivadas
8.
Cell Mol Life Sci ; 74(7): 1191-1210, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27742926

RESUMO

NDE1 (Nuclear Distribution Element 1, also known as NudE) and NDEL1 (NDE-Like 1, also known as NudEL) are the mammalian homologues of the fungus nudE gene, with important and at least partially overlapping roles for brain development. While a large number of studies describe the various properties and functions of these proteins, many do not directly compare the similarities and differences between NDE1 and NDEL1. Although sharing a high degree structural similarity and multiple common cellular roles, each protein presents several distinct features that justify their parallel but also unique functions. Notably both proteins have key binding partners in dynein, LIS1 and DISC1, which impact on neurodevelopmental and psychiatric illnesses. Both are implicated in schizophrenia through genetic and functional evidence, with NDE1 also strongly implicated in microcephaly, as well as other neurodevelopmental and psychiatric conditions through copy number variation, while NDEL1 possesses an oligopeptidase activity with a unique potential as a biomarker in schizophrenia. In this review, we aim to give a comprehensive overview of the various cellular roles of these proteins in a "bottom-up" manner, from their biochemistry and protein-protein interactions on the molecular level, up to the consequences for neuronal differentiation, and ultimately to their importance for correct cortical development, with direct consequences for the pathophysiology of neurodevelopmental and mental illness.


Assuntos
Proteínas de Transporte/genética , Proteínas Associadas aos Microtúbulos/genética , Transtornos do Neurodesenvolvimento/genética , Encéfalo/metabolismo , Proteínas de Transporte/classificação , Proteínas de Transporte/metabolismo , Pontos de Checagem do Ciclo Celular , Dineínas/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/classificação , Proteínas Associadas aos Microtúbulos/metabolismo , Transtornos do Neurodesenvolvimento/patologia , Peptídeo Hidrolases/metabolismo , Regiões Promotoras Genéticas , Esquizofrenia/genética , Esquizofrenia/patologia
9.
Proteins ; 82(7): 1210-8, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25050442

RESUMO

Carrier proteins (CPs) play a critical role in the biosynthesis of various natural products, especially in nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) enzymology, where the CPs are referred to as peptidyl-carrier proteins (PCPs) or acyl-carrier proteins (ACPs), respectively. CPs can either be a domain in large multifunctional polypeptides or standalone proteins, termed Type I and Type II, respectively. There have been many biochemical studies of the Type I PKS and NRPS CPs, and of Type II ACPs. However, recently a number of Type II PCPs have been found and biochemically characterized. In order to understand the possible interaction surfaces for combinatorial biosynthetic efforts we crystallized the first characterized and representative Type II PCP member, BlmI, from the bleomycin biosynthetic pathway from Streptomyces verticillus ATCC 15003. The structure is similar to CPs in general but most closely resembles PCPs. Comparisons with previously determined PCP structures in complex with catalytic domains reveals a common interaction surface. This surface is highly variable in charge and shape, which likely confers specificity for interactions. Previous nuclear magnetic resonance (NMR) analysis of a prototypical Type I PCP excised from the multimodular context revealed three conformational states. Comparison of the states with the structure of BlmI and other PCPs reveals that only one of the NMR states is found in other studies, suggesting the other two states may not be relevant. The state represented by the BlmI crystal structure can therefore serve as a model for both Type I and Type II PCPs.


Assuntos
Proteínas de Bactérias/química , Proteínas de Transporte/química , Modelos Moleculares , Sequência de Aminoácidos , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Biologia Computacional , Peptídeos e Proteínas de Sinalização Intracelular , Dados de Sequência Molecular , Filogenia , Conformação Proteica , Alinhamento de Sequência
10.
PLoS One ; 9(1): e84556, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24497919

RESUMO

BACKGROUND: Plant non-specific lipid transfer proteins (nsLtps) are small, basic proteins encoded by multigene families and have reported functions in many physiological processes such as mediating phospholipid transfer, defense reactions against phytopathogens, the adaptation of plants to various environmental conditions, and sexual reproduction. To date, no genome-wide overview of the Brassica rapa nsLtp (BrnsLtp) gene family has been performed. Therefore, as the first step and as a helpful strategy to elucidate the functions of BrnsLtps, a genome-wide study for this gene family is necessary. METHODOLOGY/PRINCIPAL FINDING: In this study, a total of 63 putative BrnsLtp genes were identified through a comprehensive in silico analysis of the whole genome of B. rapa. Based on the sequence similarities, these BrnsLtps was grouped into nine types (I, II, III, IV, V, VI, VIII, IX, and XI). There is no type VII nsLtps in B. rapa, and a new type, XI nsLtps, was identified in B. rapa. Furthermore, nine type II AtLtps have no homologous genes in B. rapa. Gene duplication analysis demonstrated that the conserved collinear block of each BrnsLtp is highly identical to those in Arabidopsis and that both segmental duplications and tandem duplications seem to play equal roles in the diversification of this gene family. Expression analysis indicated that 29 out of the 63 BrnsLtps showed specific expression patterns. After careful comparison and analysis, we hypothesize that some of the type I BrnsLtps may function like Arabidopsis pathogenesis-related-14 (PR-14) proteins to protect the plant from phytopathogen attack. Eleven BrnsLtps with inflorescence-specific expression may play important roles in sexual reproduction. Additionally, BrnsLtpI.3 may have functions similar to Arabidopsis LTP1. CONCLUSIONS/SIGNIFICANCE: The genome-wide identification, bioinformatic analysis and expression analysis of BrnsLtp genes should facilitate research of this gene family and polyploidy evolution and provide new insight towards elucidating their biological functions in plants.


Assuntos
Brassica rapa/genética , Proteínas de Transporte/genética , Regulação da Expressão Gênica de Plantas , Genoma de Planta/genética , Proteínas de Plantas/genética , Sequência de Aminoácidos , Sequência de Bases , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Biologia Computacional , Variação Genética , Modelos Moleculares , Dados de Sequência Molecular , Família Multigênica/genética , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/classificação , Conformação Proteica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico
11.
FEBS Lett ; 588(1): 92-8, 2014 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-24269229

RESUMO

The 14-3-3 proteins are important effectors of Ser/Thr phosphorylation in eukaryotic cells. Using mathematical modelling we investigated the roles of these proteins as effectors in signalling pathways that involve multi-phosphorylation events. We defined optimal conditions for positive and negative cross-talk. Particularly, synergistic signal interaction was evident at very different sets of binding affinities and phosphorylation kinetics. We identified three classes of 14-3-3 targets that all have two binding sites, but displayed synergistic interaction between converging signalling pathways for different ranges of parameter values. Consequently, these protein targets will respond differently to interventions that affect 14-3-3 binding affinities or phosphorylation kinetics.


Assuntos
Proteínas 14-3-3/metabolismo , Proteínas de Transporte/metabolismo , Modelos Biológicos , Transdução de Sinais , Proteínas 14-3-3/química , Sítios de Ligação , Ligação Competitiva , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Humanos , Cinética , Modelos Moleculares , Fosforilação , Ligação Proteica , Estrutura Terciária de Proteína , Proto-Oncogene Mas
12.
Mol Biol Evol ; 30(9): 2013-23, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23770652

RESUMO

Filopodia are fine actin-based cellular projections used for both environmental sensing and cell motility, and they are essential organelles for metazoan cells. In this study, we reconstruct the origin of metazoan filopodia and microvilli. We first report on the evolutionary assembly of the filopodial molecular toolkit and show that homologs of many metazoan filopodial components, including fascin and myosin X, were already present in the unicellular or colonial progenitors of metazoans. Furthermore, we find that the actin crosslinking protein fascin localizes to filopodia-like structures and microvilli in the choanoflagellate Salpingoeca rosetta. In addition, homologs of filopodial genes in the holozoan Capsaspora owczarzaki are upregulated in filopodia-bearing cells relative to those that lack them. Therefore, our findings suggest that proteins essential for metazoan filopodia and microvilli are functionally conserved in unicellular and colonial holozoans and that the last common ancestor of metazoans bore a complex and specific filopodial machinery.


Assuntos
Evolução Biológica , Coanoflagelados/classificação , Mesomycetozoea/classificação , Microvilosidades/classificação , Filogenia , Pseudópodes/classificação , Actinas/classificação , Animais , Proteínas de Transporte/classificação , Movimento Celular/fisiologia , Coanoflagelados/genética , Gelsolina/classificação , Humanos , Mesomycetozoea/genética , Proteínas dos Microfilamentos/classificação , Microvilosidades/genética , Miosinas/classificação , Pseudópodes/genética , Proteína cdc42 de Ligação ao GTP/classificação
13.
Hum Genomics ; 7: 13, 2013 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-23676014

RESUMO

The Kelch-like (KLHL) gene family encodes a group of proteins that generally possess a BTB/POZ domain, a BACK domain, and five to six Kelch motifs. BTB domains facilitate protein binding and dimerization. The BACK domain has no known function yet is of functional importance since mutations in this domain are associated with disease. Kelch domains form a tertiary structure of ß-propellers that have a role in extracellular functions, morphology, and binding to other proteins. Presently, 42 KLHL genes have been classified by the HUGO Gene Nomenclature Committee (HGNC), and they are found across multiple human chromosomes. The KLHL family is conserved throughout evolution. Phylogenetic analysis of KLHL family members suggests that it can be subdivided into three subgroups with KLHL11 as the oldest member and KLHL9 as the youngest. Several KLHL proteins bind to the E3 ligase cullin 3 and are known to be involved in ubiquitination. KLHL genes are responsible for several Mendelian diseases and have been associated with cancer. Further investigation of this family of proteins will likely provide valuable insights into basic biology and human disease.


Assuntos
Proteínas de Transporte/genética , Doenças Genéticas Inatas/genética , Família Multigênica/genética , Neoplasias/genética , Sequências Repetitivas de Aminoácidos/genética , Proteínas de Transporte/classificação , Proteínas de Transporte/metabolismo , Humanos , Neoplasias/etiologia , Filogenia , Ligação Proteica/genética , Estrutura Terciária de Proteína/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Ubiquitinação/genética
14.
Yi Chuan ; 35(3): 307-14, 2013 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-23575536

RESUMO

Plant non-specific lipid transfer proteins (nsLTPs) are a group of abundantly expressed small basic proteins, which can reversibly bind and transport hydrophobic molecules in vitro. Nine types of nsLTP have been identified from a variety of plants. All the type of nsLTPs possesses the conserved eight cysteine residue motif with a three-dimensional structure of an internal hydrophobic cavity and the lipid binding site. Based on the growing knowledge about the structure, gene expression, regulation and in vitro activity, nsLTPs are considered to play a role in key processes of plant physiology, including wax synthesis and transport, abiotic stress resistance, disease resistance, and plant reproduction. This review aims at presenting comprehensive information of key topics, including basic features, classification, gene cloning, expression profiles, and functional studies of nsLTP. Finally the perspectives were included on the future study of nsLTP family.


Assuntos
Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Plantas/genética , Plantas/metabolismo , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/química , Proteínas de Plantas/classificação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
15.
J Alzheimers Dis ; 29(4): 863-73, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22349685

RESUMO

Zinc (Zn2+) is concentrated into pre-synaptic vesicles and co-released with neurotransmitter at some synapses. Zn2+ can accelerate assembly of the amyloid-ß peptides (Aß) and tau protein central to the neuropathological changes found in Alzheimer's disease (AD). Altered protein levels of the membrane Zn2+ transporters ZnT1, ZnT4, and ZnT6 have been reported in AD postmortem brain tissue. The present study analyzed mRNA levels of five established (LIV1, ZIP1, ZnT1, ZnT4, and ZnT6) and one potential (PRNP) Zn2+ transporter in human postmortem brain tissue from Braak-staged individuals with AD and controls using quantitative real-time PCR. Four cortical regions (middle temporal gyrus, superior occipital gyrus, superior parietal gyrus, and superior frontal gyrus) and cerebellum were examined. PRNP mRNA levels were decreased by ∼30% in all four cortical regions examined in AD patients, but unchanged in the cerebellum. In contrast, some increases in mRNA levels of the other more established Zn2+ transporters (LIV1, ZIP1, ZnT1, ZnT6) were found in AD cortex. The ratios of the mRNA levels of LIV1, ZIP1, ZnT1, ZnT4, and ZnT6/mRNA level of neuron specific enolase increased significantly as the disease progressed and Braak stage increased. Significant correlations were also identified between mRNA levels of several of the Zn2+ transporters investigated. These expression changes could either reflect or cause the altered cortical Zn2+ distribution in AD, potentially increasing the likelihood of interactions between Zn2+ and Aß or tau protein.


Assuntos
Doença de Alzheimer/patologia , Encéfalo/metabolismo , Proteínas de Transporte/genética , RNA Mensageiro/metabolismo , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Encéfalo/patologia , Proteínas de Transporte/classificação , Proteínas de Transporte/metabolismo , Estudos de Casos e Controles , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Feminino , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Masculino , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Mudanças Depois da Morte , Estatística como Assunto , Estatísticas não Paramétricas
16.
J Proteome Res ; 9(12): 6298-316, 2010 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-20873783

RESUMO

Physical protein-protein interactions are fundamental to all biological processes and are organized in complex networks. One branch of the kinome network is the evolutionarily conserved NIMA-related serine/threonine kinases (Neks). Most of the 11 mammalian Neks studied so far are related to cell cycle regulation, and due to association with diverse human pathologies, Neks are promising chemotherapeutic targets. Human Nek6 was associated to carcinogenesis, but its interacting partners and signaling pathways remain elusive. Here we introduce hNek6 as a highly connected member in the human kinase interactome. In a more global context, we performed a broad data bank comparison based on degree distribution analysis and found that the human kinome is enriched in hubs. Our networks include a broad set of novel hNek6 interactors as identified by our yeast two-hybrid screens classified into 18 functional categories. All of the tested interactions were confirmed, and the majority of tested substrates were phosphorylated in vitro by hNek6. Notably, we found that hNek6 N-terminal is important to mediate the interactions with its partners. Some novel interactors also colocalized with hNek6 and γ-tubulin in human cells, pointing to a possible centrosomal interaction. The interacting proteins link hNek6 to novel pathways, for example, Notch signaling and actin cytoskeleton regulation, or give new insights on how hNek6 may regulate previously proposed pathways such as cell cycle regulation, DNA repair response, and NF-κB signaling. Our findings open new perspectives in the study of hNek6 role in cancer by analyzing its novel interactions in specific pathways in tumor cells, which may provide important implications for drug design and cancer therapy.


Assuntos
Proteínas de Transporte/metabolismo , Centrossomo/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteômica/métodos , Sítios de Ligação , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Células HEK293 , Humanos , Microscopia Confocal , Modelos Biológicos , Quinases Relacionadas a NIMA , Ligação Proteica , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Técnicas do Sistema de Duplo-Híbrido
17.
J Exp Bot ; 61(15): 4277-90, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20667964

RESUMO

Lily stigma/style cysteine-rich adhesin (SCA), a plant lipid transfer protein (LTP) which is secreted into the extracellular matrix, functions in pollen tube guidance in fertilization. A gain-of-function mutant (ltp5-1) for Arabidopsis LTP5, an SCA-like molecule, was recently shown to display defects in sexual reproduction. In the current study, it is reported that ltp5-1 plants have dwarfed primary shoots, delayed hypocotyl elongation, various abnormal tissue fusions, and display multibranching. These mutant phenotypes in vegetative growth are recessive. No abnormality was found in ltp5-1/+ plants. In a phylogenetic analysis of plant LTPs, SCA-like Arabidopsis LTPs were classified with conventional plant LTPs. Homology modelling-based electrostatic similarity index (ESI) clustering was used to show diversity in spatial distributions of electrostatic potentials of SCA-like LTPs, suggestive of their various roles in interaction in the extracellular matrix space. ß-Glucuronidase (GUS) analysis showed that SCA-like Arabidopsis LTP genes are diversely present in various tissues. LTP4 was found specifically in the guard cells and LTP6 in trichomes as well as in other tissues. LTP1 levels were specifically abundant in the stigma, and both LTP3 and LTP6 in the ovules. LTP2 and LTP4 gene levels were up-regulated in whole seedlings with 20% polyethylene glycol (PEG) and 300 mM NaCl treatments, respectively. LTP5 was up-regulated in the hypocotyl with 3 d dark growth conditions. LTP6 was specifically expressed in the tip of the cotyledon under drought stress conditions. The results suggest that SCA-like Arabidopsis LTPs are multifunctional, with diversified roles in plant growth and reproduction.


Assuntos
Antígenos de Plantas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Transporte/metabolismo , Cisteína/metabolismo , Flores/metabolismo , Proteínas de Plantas/metabolismo , Antígenos de Plantas/química , Antígenos de Plantas/classificação , Antígenos de Plantas/genética , Arabidopsis/citologia , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/classificação , Proteínas de Arabidopsis/genética , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Análise por Conglomerados , Flores/citologia , Flores/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas/genética , Mutação/genética , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/classificação , Proteínas de Plantas/genética , Reprodução/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Estresse Fisiológico/genética , Homologia Estrutural de Proteína
18.
J Proteome Res ; 9(6): 3311-8, 2010 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-20405931

RESUMO

A straightforward method using mild enzymatic digestions combined with MALDI mass spectrometry (MS) was used to enhance determination of the multiple phosphorylation sites of a set of recombinant nucleotide-binding proteins in Escherichia coli, including kinases and cystathionine beta-synthase (CBS) domain containing proteins. The protein kinases reveal abundant phosphorylations in the kinase domains and relatively low phosphogluconoylation (258 Da) at the N-terminal His-tag. In contrast, the CBS domain-containing proteins possess a highly conserved phosphorylation in vivo at Ser-2 of the His-tag. Multistage MS/MS and selected reaction monitoring established that the CBS domain proteins also contain a combined modification of gluconoylation (178 Da) and phosphorylation (80 Da) at two different sites, instead of an isobaric phosphogluconoylation (258 Da) event at the N-terminus. Functional analysis of 20 recombinant proteins as identified by mass spectrometry has shown the phosphorylation at the N-terminal His-tag is relevant to nucleotide binding and phosphotransfer reaction catalyzed by a serine protein kinase.


Assuntos
Escherichia coli/química , Fragmentos de Peptídeos/química , Proteínas Recombinantes de Fusão/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Sequência de Aminoácidos , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Proteínas de Transporte/metabolismo , Escherichia coli/metabolismo , Histidina/química , Dados de Sequência Molecular , Oligopeptídeos/química , Fragmentos de Peptídeos/metabolismo , Fosforilação , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/classificação , Proteínas Recombinantes de Fusão/metabolismo , Espectrometria de Massas em Tandem , Tripsina/metabolismo
19.
Liver Int ; 30(3): 388-95, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19968777

RESUMO

BACKGROUND/AIMS: Transcription factors coupled to cyclic adenosine mono phosphate (cAMP) signalling in the cAMP-responsive elements binding (CREB)/ATF family constitute a family of activators or repressors that bind to cAMP-responsive promoter elements (CREs) in the regulatory regions of cAMP-inducible genes. A role for CREB/ATF family has been advocated in the control of hepatocellular carcinoma progression. CREB appears to be activated by the X protein of hepatitis B virus, which links to the unphosphorylated form of CREB and activates transcription, thus obviating an otherwise indispensable Ser-133 phosphorylation. Identification of factors capable of triggering transcription via cAMP-responsive elements modulator (CREM)/CREB signalling in the absence of Ser phosphorylation will improve our knowledge of the molecular mechanism of liver cell proliferation. METHODS: To isolate and study proteins binding and activating CREB and/or CREM in the liver, we performed the screening of a mouse liver cDNA library using the Two-Hybrid System. RESULTS: We report the identification and characterization of a novel peptide, VTIP-peptide (VTIP-P), which binds and enhances the activation of CREM/CREB, obviating the need for transcription factor phosphorylation. We demonstrated that VTIP-P physically interacts with the activation domain (AD) of the transcription factors CREB/CREM and activates transcription by modifying their phosphorylation pattern in hepatoma cells. The data allowed the conclusion that VTIP-P binds the AD of CREB and CREM by stabilizing their phosphorylation. CONCLUSION: The characterization of molecules capable of interfering in the liver with an important pathway such as CREB could be significant in designing and/or developing new therapeutic approaches to the control of liver cell proliferation.


Assuntos
Proteínas de Transporte/metabolismo , Modulador de Elemento de Resposta do AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regeneração Hepática/fisiologia , Fígado/metabolismo , Peptídeos/metabolismo , Animais , Proteínas de Transporte/classificação , Linhagem Celular Tumoral , Modulador de Elemento de Resposta do AMP Cíclico/genética , Biblioteca Gênica , Camundongos , Peptídeos/classificação , Fatores de Transcrição/metabolismo , Transfecção , Técnicas do Sistema de Duplo-Híbrido
20.
Dev Cell ; 17(1): 110-22, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19619496

RESUMO

Early endosome-to-trans-Golgi network (TGN) transport is organized by the retromer complex. Consisting of cargo-selective and membrane-bound subcomplexes, retromer coordinates sorting with membrane deformation and carrier formation. Here, we describe four mammalian retromers whose membrane-bound subcomplexes contain specific combinations of the sorting nexins (SNX), SNX1, SNX2, SNX5, and SNX6. We establish that retromer requires a dynamic spatial organization of the endosomal network, which is regulated through association of SNX5/SNX6 with the p150(glued) component of dynactin, an activator of the minus-end directed microtubule motor dynein; an association further defined through genetic studies in C. elegans. Finally, we also establish that the spatial organization of the retromer pathway is mediated through the association of SNX1 with the proposed TGN-localized tether Rab6-interacting protein-1. These interactions describe fundamental steps in retromer-mediated transport and establish that the spatial organization of the retromer network is a critical element required for efficient retromer-mediated sorting.


Assuntos
Proteínas de Transporte/metabolismo , Dineínas/metabolismo , Endossomos/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Rede trans-Golgi/metabolismo , Animais , Transporte Biológico/fisiologia , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Linhagem Celular , Complexo Dinactina , Dineínas/genética , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Complexos Multiproteicos/metabolismo , Filogenia , Isoformas de Proteínas/genética , Interferência de RNA , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Nexinas de Classificação , Técnicas do Sistema de Duplo-Híbrido , Proteínas de Transporte Vesicular/classificação , Proteínas de Transporte Vesicular/genética , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
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