RESUMO
Contryphans, peptides containing a single disulfide bond, are found abundantly in cone snail venom. The analysis of a large dataset of available contryphan sequences permits a classification based on the occurrence of proline residues at positions 2 and 5 within the macrocyclic 23-membered disulfide loop. Further sequence diversity is generated by variable proteolytic processing of the contryphan precursor proteins. In the majority of contryphans, presence of Pro at position 2 and a D-residue at position 3 leads to a slow conformational dynamics, manifesting as anomalous chromatographic profiles during LC analysis. LC-MS analysis of diverse contryphans suggests that elution profiles may be used as a rapid diagnostic for the presence of the Pro2-DXxx3 motif. Natural sequences from C.inscriptus and C.frigidus together with synthetic analogs permit the delineation of the features necessary for abnormal chromatographic behaviour. A diagnostic for the presence of Pro at position 5 is obtained by the observation of non-canonical fragment ions, generated by N-Cα bond cleavage at the dehydroalanine residue formed by disulfide cleavage. Anomalous LC profiles supports Pro at position 2, while non-canonical mass spectral fragments established Pro at position 5, providing a rapid method for contryphan analysis from LC-ESI-MS/MS profiles of crude Conus venom. SIGNIFICANCE: Contryphans are peptides, widely distributed in cone snail venom, which display extensive sequence diversity. Heterogeneity of proteolytic processing of contryphan precursor proteins, together with post-translational modifications contributes to contryphan diversity. Contryphans, identified by a combination of mass spectrometry and transcriptomic analysis, are classified on the basis of sequence features, primarily the number of proline residues within the disulfide loop. Conformational diversity arises in contryphans by cis-trans isomerization of Cys-Pro bonds, resulting in characteristic chromatographic profiles, permitting identification even in crude venom mixtures. Rapid identification of contryphans in cone snail peptide libraries is also facilitated by diagnostic mass spectral fragments arising by non-canonical cleavage of the N-Cα bond at Cys(7).
Assuntos
Conotoxinas , Caramujo Conus , Animais , Espectrometria de Massas em Tandem , Sequência de Aminoácidos , Peptídeos/química , Venenos de Moluscos/química , Dissulfetos/química , Prolina , Caramujo Conus/química , Conotoxinas/químicaRESUMO
AIM: To understand the mechanism of glycation of albumin and effects on cysteinylation and methionine oxidation. METHODS: The in vitro glycation of HSA and BSA was studied with varying concentrations of glucose. Clinical blood samples of diabetic subjects with varying HbA1c values, were analyzed to assess in vivo glycation. All samples and their tryptic digests were analyzed using liquid chromatography/mass spectrometry. Glycation sites were mapped on to the three-dimensional structure of the HSA and BSA. RESULTS: A total thirty-one sites for glycation and eight sites of Nε-carboxymethyl-lysine (CML) modification were identified on albumin. The site selectivity of glycation was correlated with the environment of the reactive residue in the three-dimensional structure. CONCLUSIONS: The maximum percentage glycation under extreme conditions was in the range of ~55 to 88% in four weeks. Two major glycation sites K-233 and K-525 were identified, which together accounted for 40-50% of total glycation. A correlation was observed between glycation and oxidation of methionine residues in samples glycated in vitro. The role of spatially proximate residues in facilitating the glycation process is evident. The tri- and tetra-glycated isoforms of albumin can serve as biomarkers for the severe uncontrolled diabetic state.
Assuntos
Diabetes Mellitus , Albumina Sérica , Glucose , Produtos Finais de Glicação Avançada , Glicosilação , Humanos , Espectrometria de Massas , Albumina Sérica/metabolismoRESUMO
Four 30 residue conotoxin have been identified from the venom of C. amadis. MS/MS analysis of crude venom subjected to global reduction/alkylation yielded fragmentation patterns, which permitted searching and matching with a database of putative mature toxin sequences obtained from transcriptomic analysis. Of the four sequences identified, Am3408(Am6.1b), Am3452(Am6.1c), Am3136(Am6.2a) and Am3214(Am6.2b), three contain bromotryptophan residues, while an additional post translational modification, gamma carboxylation of glutamic acid, is present in Am3408(Am6.1b)/3452(Am6.1c). The conotoxins belong to the O1/O2 gene superfamily and possess cysteine framework VI/VII. While, the cysteine patterns show a similarity to omega conotoxins, the three C. amadis peptides are highly negatively charged and possess a significant content of hydrophobic residues.
Assuntos
Conotoxinas/química , Peptídeos/química , Sequência de Aminoácidos , Animais , Caramujo Conus/química , Peptídeos/isolamento & purificação , Processamento de Proteína Pós-Traducional , Espectrometria de Massas em Tandem , Transcriptoma , Triptofano/químicaRESUMO
Lung cancer, the most common cause of cancer-related death in men and women, is responsible for 1.3 million deaths worldwide annually. Women are diagnosed to a greater extent than men with adenocarcinoma and small cell carcinoma, both of which are secretory-type tumors. Never smokers diagnosed with lung cancer are also predominantly female, demonstrating the association of genetic factors with lung carcinogenesis. Several epidemiologic studies have associates certain CYP1A1 genotypes, alone or in combination, with an increased risk of estrogen-related cancer. The aim of this study was to investigate the impact of the CYP and GST polymorphisms along with estrogen and interleukin-6 (IL-6) levels on the risk of lung cancer. Eighty-six lung cancer patients and 60 controls were included in the study. A significantly higher frequency of polymorphisms in the genes was observed in lung cancer patients compared to controls. Mean estradiol concentration was reduced and IL-6 levels were elevated in patients compared to controls. In conclusion, increased polymorphisms in metabolic genes may be the reason for the reduced estradiol and, thereby, the increased expression of IL-6 in the serum of lung cancer patients.
RESUMO
Dimeric and monomeric forms of the enzyme triosephosphate isomerase (TIM) from Plasmodium falciparum (Pf) have been detected under conditions of nanoflow by electrospray mass spectrometry. The dimer (M = 55 663 Da) exhibits a narrow charge state distribution with intense peaks limited to values of 18(+) to 21(+), maximal intensity being observed for charge states 19(+) and 20(+). A monomeric species with a charge state distribution ranging from 11(+) to 16(+) is also observed, which may be assigned to folded dissociated subunits. Complete dimer dissociation results under normal electrospray condition. The effects of solution pH and source temperature have been investigated. The observation of four distinct charge state distributions which may be assigned to a dimer, folded monomer, partially folded monomer and unfolded monomer is reported. Circular dichromism and fluorescence studies of Pf TIM at low pH support the retention of substantial secondary and tertiary structures. Satellite peaks in mass spectra corresponding to hydrated species are also observed and isotope shift upon deuteration is demonstrated. The analysis of all available independent crystal structures of Pf TIM and TIMs from other organisms permits identification of structurally conserved water molecules. Hydration observed in the dimer and folded monomeric forms in the gas phase may correspond to these conserved sites.
Assuntos
Modelos Moleculares , Plasmodium falciparum/enzimologia , Triose-Fosfato Isomerase/genética , Triose-Fosfato Isomerase/metabolismo , Animais , Dicroísmo Circular , Cristalografia , Dimerização , Concentração de Íons de Hidrogênio , Isoenzimas/genética , Espectrometria de Massas , Plasmodium falciparum/genética , Conformação Proteica , TemperaturaRESUMO
A synthetic collagenase substrate containing the internal peptide sequence--Gly-Gly-Pro-Leu-Gly-Pro-Pro-Gly-Pro--has been synthesized, with an N-terminus 4-((4-(dimethylamino)phenyl)azo)-benzoyl (DABCYL) group and C-terminus 5-[2-(acetamido)ethylamino] naphthalene-1-sulfonic acid (AEDANS) moiety resulting in internal quenching of AEDANS fluorescence. Peptide bond hydrolysis results in a large increase in fluorescence at 490 nm upon excitation at 336 nm. The substrate is cleaved exclusively by Clostridium histolyticum collagenase and is completely resistant to attack by proteases like thermolysin, proteinase K, and trypsin. K(m) and V(max) values for substrate hydrolysis by collagenase have been determined, establishing the peptide as one of the best binding substrates for the enzyme. MALDI mass spectrometry using a derivative of the substrate establishes that the sites of cleavage lie within the collagen like domain. The CD spectrum of an analog peptide lacking the donor and acceptor groups reveals spectral features that are reminiscent of weak polyproline structures.
Assuntos
Colagenases/metabolismo , Corantes Fluorescentes/química , Corantes Fluorescentes/metabolismo , Dicroísmo Circular , Clostridium histolyticum/enzimologia , Cinética , Dados de Sequência Molecular , Estrutura Molecular , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Especificidade por SubstratoRESUMO
Half a century has passed since the hydrogen-bonded secondary structures of polypeptides and proteins were first recognized. An extraordinary wealth of conformational information is now available on peptides and proteins, which are formed of alpha-amino acid residues. More recently, the discovery of well-folded structures in oligopeptides containing beta-amino acids has focused a great deal of current interest on the conformational properties of peptides constructed from higher homologues (omega) of alpha-amino acids. This review examines the nature of intramolecularly hydrogen-bonded conformations of hybrid peptides formed by amino acid residues, with a varying number of backbone atoms. The beta-turn, a ubiquitous structural feature formed by two residue (alphaalpha) segments in proteins and peptides, is stabilized by a 10-atom (C10) intramolecular 4-->1 hydrogen bond. Hybrid turns may be classified by comparison with their alphaalpha counterparts. The available crystallographic information on hydrogen-bonded hybrid turns is surveyed in this review. Several recent examples demonstrate that individual omega-amino acid residues and hybrid dipeptide segments may be incorporated into the regular structures of alpha-peptides. Examples of both peptide helices and hairpins are presented. The present review explores the relationships between folded conformations in hybrid sequences and their counterparts in all alpha-residue sequences. The use of stereochemically constrained omega-residues promises to expand the range of peptide design strategies to include omega-amino acids. This approach is exemplified by well-folded structures like the C12 (alphagamma) and C14 (gammagamma) helices formed in short peptides containing multiply substituted gamma-residues. The achiral gamma-residue gabapentin is a readily accessible building block in the design of peptides containing gamma-amino acids. The construction of globular polypeptide structures using diverse hybrid sequences appears to be a realistic possibility.
Assuntos
Aminoácidos/química , Peptídeos/química , Ligação de Hidrogênio , Isomerismo , Estrutura Secundária de ProteínaRESUMO
Distinctly different effects of two closely related contryphans have been demonstrated on voltage-activated Ca(2+) channels. The peptides Lo959 and Am975 were isolated from Conus loroisii, a vermivorous marine snail and Conus amadis, a molluscivore, respectively. The sequences of Lo959 and Am975 were deduced by mass spectrometric sequencing (MALDI-MS/MS) and confirmed by chemical synthesis. The sequences of Lo959, GCP(D)WDPWC-NH(2) and Am975, GCO(D)WDPWC-NH(2) (O: 4-trans-hydroxyproline: Hyp), differ only at residue 3; Pro in Lo959, Hyp in Am975, which is identical to contryphan-P, previously isolated from Conus purpurascens, a piscivore; while Lo959 is a novel peptide. Both Lo959 and Am975 undergo slow conformational interconversion under reverse-phase chromatographic conditions, a characteristic feature of all contryphans reported thus far. Electrophysiological studies performed using dorsal root ganglion neurons reveal that both peptides target high voltage-activated Ca(2+) channels. While Lo959 increases the Ca(2+) current, Am975 causes inhibition. The results establish that subtle sequence effects, which accompany post-translational modifications in Conus peptides, can have dramatic effects on target ion channels.
Assuntos
Canais de Cálcio/fisiologia , Conotoxinas/química , Caramujo Conus/química , Peptídeos Cíclicos/química , Peptídeos/química , Peptídeos/genética , Sequência de Aminoácidos , Animais , Canais de Cálcio/efeitos dos fármacos , Células Cultivadas , Dados de Sequência Molecular , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Técnicas de Patch-Clamp , Peptídeos Cíclicos/farmacologia , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Two designed peptide sequences containing Trp residues at positions i and i + 5 (Boc-Leu-Trp-Val-Ala-Aib-Leu-Trp-Val-OMe, 1) as well as i and i + 6 (Boc-Leu-Trp-Val-Aib-Ala-Aib-Leu-Trp-Val-OMe, 2) containing one and two centrally positioned Aib residues, respectively, for helix nucleation, have been shown to form stable helices in chloroform solutions. Structures derived from nuclear magnetic resonance (NMR) data reveal six and seven intramolecularly hydrogen-bonded NH groups in peptides 1 and 2, respectively. The helical conformation of octapeptide 1 has also been established in the solid state by X-ray diffraction. The crystal structure reveals an interesting packing motif in which helical columns are stabilized by side chain-backbone hydrogen bonding involving the indole Nepsilon1H of Trp(2) as donor, and an acceptor C=O group from Leu(6) of a neighboring molecule. Helical columns also associate laterally, and strong interactions are observed between the Trp(2) and Trp(7) residues on neighboring molecules. The edge-to-face aromatic interactions between the indoles suggest a potential C-H...pi interaction involving the Czeta3H of Trp(2). Concentration dependence of NMR chemical shifts provides evidence for peptide association in solution involving the Trp(2) Nepsilon1H protons, presumably in a manner similar to that observed in the crystal.
Assuntos
Indóis/metabolismo , Peptídeos/química , Triptofano/química , Cristalografia por Raios X , Ligação de Hidrogênio , Indóis/química , Modelos Moleculares , Conformação Molecular , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Triptofano/metabolismoRESUMO
The crystal structures of the peptides, Boc-Leu-Trp-Val-OMe (1), Ac-Leu-Trp-Val-OMe (2a and 2b), Boc-Leu-Phe-Val-OMe (3), Ac-Leu-Phe-Val-OMe (4), and Boc-Ala-Aib-Leu-Trp-Val-OMe (5) have been determined by X-ray diffraction in order to explore the nature of interactions between aromatic rings, specifically the indole side chain of Trp residues. Peptide 1 adopts a type I beta-turn conformation stabilized by an intramolecular 4-->1 hydrogen bond. Molecules of 1 pack into helical columns stabilized by two intermolecular hydrogen bonds, Leu(1)NH...O(2)Trp(2) and IndoleNH...O(1)Leu(1). The superhelical columns further pack into the tetragonal space group P4(3) by means of a continuous network of indole-indole interactions. Peptide 2 crystallizes in two polymorphic forms, P2(1) (2a) and P2(1)2(1)2(1) (2b). In both forms, the peptide backbone is extended, with antiparallel beta-sheet association being observed in crystals. Extended strand conformations and antiparallel beta-sheet formation are also observed in the Phe-containing analogs, Boc-Leu-Phe-Val-OMe (3) and Ac-Leu-Phe-Val-OMe (4). Peptide 5 forms a short stretch of 3(10)-helix. Analysis of aromatic-aromatic and aromatic-amide interactions in the structures of peptides, 1, 2a, 2b are reported along with the examples of 14 Trp-containing peptides from the Cambridge Crystallographic Database. The results suggest that there is no dramatic preference for a preferred orientation of two proximal indole rings. In Trp-containing peptides specific orientations of the indole ring, with respect to the preceding and succeeding peptide units, appear to be preferred in beta-turns and extended structures.
Assuntos
Peptídeos/química , Fenilalanina/química , Triptofano/química , Cristalização , Cristalografia por Raios X , Ligação de Hidrogênio , Modelos Moleculares , Conformação ProteicaRESUMO
Highly structured small peptides are the major toxic constituents of the venom of cone snails, a family of widely distributed predatory marine molluscs. These animals use the venom for rapid prey immobilization. The peptide components in the venom target a wide variety of membrane-bound ion channels and receptors. Many have been found to be highly selective for a diverse range of mammalian ion channels and receptors associated with pain-signaling pathways. Their small size, structural stability, and target specificity make them attractive pharmacologic agents. A select number of laboratories mainly from the United States, Europe, Australia, Israel, and China have been engaged in intense drug discovery programs based on peptides from a few snail species. Coastal India has an estimated 20-30% of the known cone species; however, few serious studies have been reported so far. We have begun a comprehensive program for the identification and characterization of peptides from cone snails found in Indian Coastal waters. This presentation reviews our progress over the last 2 years. As expected from the evolutionary history of these venom components, our search has yielded novel peptides of therapeutic promise from the new species that we have studied.
Assuntos
Produtos Biológicos/uso terapêutico , Peptídeos/isolamento & purificação , Peptídeos/uso terapêutico , Sequência de Aminoácidos , Animais , Índia , Canais Iônicos/efeitos dos fármacos , Canais Iônicos/fisiologia , Moluscos , Venenos de Moluscos/uso terapêutico , Peptídeos Cíclicos/química , Peptídeos Cíclicos/isolamento & purificaçãoRESUMO
Conformational studies on the synthetic 11-aa peptide t-butoxycarbonyl (Boc)-Val-Ala-Phe-alpha-aminoisobutyric acid (Aib)-(R)-beta3-homovaline (betaVal)-(S)-beta3-homophenylalanine (betaPhe)-Aib-Val-Ala-Phe-Aib-methyl ester (OMe) (peptide 1; betaVal and betaPhe are beta amino acids generated by homologation of the corresponding l-residues) establish that insertion of two consecutive beta residues into a polypeptide helix can be accomplished without significant structural distortion. Crystal-structure analysis reveals a continuous helical conformation encompassing the segment of residues 2-10 of peptide 1. At the site of insertion of the betabeta segment, helical hydrogen-bonded rings are expanded. A C15 hydrogen bond for the alphabetabeta segment and two C14 hydrogen bonds for the alphaalphabeta or betaalphaalpha segments have been characterized. The following conformational angles were determined from the crystal structure for the beta residues: betaVal-5 (= -126 degrees, = 76 degrees, and psi = -124) and betaPhe-6 (=-88 degrees, = 80 degrees, and psi =-118). The N terminus of the peptide is partially unfolded in crystals. The 500-MHz 1H-NMR studies establish a continuous helix over the entire length of the peptide in CDCl3 solution, as evidenced by diagnostic nuclear Overhauser effects. The presence of seven intramolecular hydrogen bonds is also established by using solvent dependence of NH chemical shifts.
Assuntos
Oligopeptídeos/química , Aminoácidos/química , Fenômenos Biofísicos , Biofísica , Cristalografia por Raios X , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Estrutura Secundária de ProteínaRESUMO
Cell adhesion has an essential role in regulating metastasis, and loss of cell adhesion is a classic feature of invasion. There is currently a great deal of interest in the role of adhesion proteins and the antimetastatic protein nm23H1 in the progression of cancer. However, reports on the expression of these proteins in complete hydatidiform moles (CHMs) are limited. In the present study, expression of the adhesion molecules E-cadherin, P-cadherin, CD44, and CD44v6, and the antimetastatic protein nm23H1 was assessed in relation to the invasive potential of CHMs. This is the first report on the expression of these proteins in CHMs. Immunohistochemical assessment was carried out in CHMs (105 cases including 15 cases of invasive moles) and compared with that of gestational age-matched normal placentae (95 cases). The expression of the adhesion proteins ranged from mild to moderate intensity with a general down-regulation in the molar trophoblasts, the down-regulation in CD44 and CD44V6 being highly significant. No relation was, however, noticed with the invasiveness or the persistence of the disease. nm23H1 protein, on the other hand, was significantly down-regulated in the molar trophoblasts with none of the invasive lesions showing intense expression. The study thus suggests down-regulation of adhesion proteins, especially that of CD44 and CD44v6, to be an early step in the transformation to molar placenta with reduced expression of nm23H1 conferring an invasive potential to the trophoblasts.
Assuntos
Moléculas de Adesão Celular/metabolismo , Mola Hidatiforme/metabolismo , Núcleosídeo-Difosfato Quinase , Neoplasias Uterinas/metabolismo , Caderinas/metabolismo , Estudos de Casos e Controles , Feminino , Glicoproteínas/metabolismo , Humanos , Receptores de Hialuronatos/metabolismo , Mola Hidatiforme/patologia , Nucleosídeo NM23 Difosfato Quinases , Invasividade Neoplásica , Gravidez , Trimestres da Gravidez , Proteínas/metabolismo , Neoplasias Uterinas/patologiaRESUMO
This review briefly surveys the conformational properties of guest omega-amino acid residues when incorporated into host alpha-peptide sequences. The results presented focus primarily on the use of beta- and gamma-residues in alphaomega sequences. The insertion of additional methylene groups into peptide backbones enhances the range of accessible conformations, introducing additional torsional variables. A nomenclature system, which permits ready comparisons between alpha-peptides and hybrid sequences, is defined. Crystal structure determination of hybrid peptides, which adopt helical and beta-hairpin conformations permits the characterization of backbone conformational parameters for beta- and gamma-residues inserted into regular alpha-polypeptide structures. Substituted beta- and gamma-residues are more limited in the range of accessible conformation than their unsubstituted counterparts. The achiral beta,beta-disubstituted gamma-amino acid, gabapentin, is an example of a stereochemically constrained residue in which the torsion angles about the Cbeta-Cgamma (theta1) and Calpha-Cbeta (theta2) bonds are restricted to the gauche conformation. Hybrid sequences permit the design of novel hydrogen bonded rings in peptide structures.
Assuntos
Aminoácidos/química , Peptídeos/química , Estrutura Molecular , Conformação ProteicaRESUMO
The aim of this study was to see whether serial cytological evaluation of various cellular abnormalities in tumours from patients receiving fractionated radiotherapy can predict radio-response in oral carcinoma. Cytological assessment was carried out in scrape smears collected prior to and during the course of radiotherapy in 68 patients with squamous cell carcinoma of the oral cavity planned for radical radiotherapy with accelerated fraction schedule. Smears were evaluated for a set of 15 radiation-induced cellular abnormalities. The relationship between the cellular alterations and the cumulative radiation dose was analysed by Kruskal-Wallis one-way anova. The results showed that among the various quantifiable changes that occur in irradiated cancer cells, karyolysis, karyorrhexis, pyknosis, cytolysis, multinucleation, micronucleation and nuclear budding show significant increase depending on the dose of radiation. The radio-resistant group of patients exhibited a lesser degree of change compared with the radio-sensitive group. This suggests that radio-resistance may be due to the defective induction of cell damage and that these cytological features may have potential use as predictive markers of radio-sensitivity in oral carcinoma.
Assuntos
Carcinoma de Células Escamosas/radioterapia , Citodiagnóstico/métodos , Neoplasias Bucais/radioterapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/patologia , Núcleo Celular/efeitos da radiação , Citoplasma/efeitos da radiação , Relação Dose-Resposta à Radiação , Humanos , Pessoa de Meia-Idade , Neoplasias Bucais/patologia , Tolerância a Radiação , Resultado do TratamentoRESUMO
Between 1996 and 1999, we carried out a study in Southern India on risk factors for oral cancer. The study included 591 incident cases of cancer of the oral cavity (282 women) and 582 hospital controls (290 women). Height was unrelated to oral cancer risk. Body mass index (weight in kilograms/height in metres squared) was inversely associated with risk (P for trend<0.001). Paan chewers with low BMI were at particularly high risk. Risk was increased among subjects consuming meat (odds ratio (OR) 1.54, 95% confidence interval (CI) 1.00-2.37), ham and salami (OR 4.40, 95% CI 2.88-6.71) two or more times per week. Frequent consumption of fish, eggs, raw green vegetables, cruciferous vegetables, carrots, pulses, apples or pears, citrus fruit, and overall consumption of vegetables and fruit decreased oral cancer risk (P for trend for each of these items less than or equal to 0.001). The risk associated with low consumption of vegetables was higher among smokers than among non-smokers. Men, but not women, who practised oral sex had an increased oral cancer risk (OR 3.14, 95% CI 1.15-8.63). Women with more than one sexual partner during life were at increased oral cancer risk (OR 9.93, 95% CI 1.57-62.9).
Assuntos
Neoplasias Bucais/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Índice de Massa Corporal , Estudos de Casos e Controles , Dieta , Feminino , Humanos , Incidência , Índia/epidemiologia , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/etiologia , Neoplasias Bucais/patologia , Fatores de Risco , Comportamento Sexual , Fumar , Inquéritos e Questionários , VerdurasRESUMO
The possible occurrence of a novel helix terminating structural motif in proteins involving a stabilizing short C-H...O interaction has been examined using a dataset of 634 non-homologous protein structures (Assuntos
Estrutura Secundária de Proteína
, Proteínas/química
, Proteínas de Bactérias/química
, Proteínas de Escherichia coli
, Conformação Proteica
, Estrutura Terciária de Proteína
RESUMO
The prognostic significance of nm23H1 and p53 proteins as predictors of nodal involvement and distant recurrence was evaluated in 63 cancer and 47 benign lesions of the breast. Assessment was carried out by immunohistochemical staining using nm23H1 and p53 antibodies. Results show no relation of nm23H1 either to the malignant nature when compared to benign lesions or to the nodal status. P53 protein, on the other hand, showed significantly increased expression in malignant lesions (p=0.001) and correlated well with nodal positivity (p=0.03). A follow-up study of 5 years showed that among the cases showing recurrence, those with positive mn23H1 showed a shorter distant recurrence free survival (DRFS, p=0.01), while p53 expression had no effect. This result did not agree with the previous reports showing nm23H1 as an antimetastatic gene. This is the first report of positive correlation between nm231-11 and distant recurrence in breast cancers, though such a correlation was reported earlier in certain other cancers. Since nm23H1 is an NDP kinase, having more involvement in signal transduction of cell proliferation, it is not difficult to comprehend such a role for nm23H1 during recurrence. Combining the expression of both proteins, lesions positive for both p53 and nm23H1 had significantly reduced distant recurrence free survival (DRFS), when compared to those negative for both proteins (p=0.0006). It can be concluded that nni23H1 alteration has a potential in predicting DRFS, while p53 alteration has the potential to predict lymph node involvement.
Assuntos
Neoplasias da Mama/diagnóstico , Carcinoma/diagnóstico , Proteínas Monoméricas de Ligação ao GTP/análise , Núcleosídeo-Difosfato Quinase , Fatores de Transcrição/análise , Proteína Supressora de Tumor p53/análise , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Carcinoma/mortalidade , Carcinoma/secundário , Intervalo Livre de Doença , Feminino , Humanos , Imuno-Histoquímica , Metástase Linfática , Proteínas Monoméricas de Ligação ao GTP/imunologia , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Nucleosídeo NM23 Difosfato Quinases , Metástase Neoplásica , Prognóstico , Recidiva , Análise de Sobrevida , Fatores de Transcrição/imunologia , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/imunologia , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/análise , Proteínas Supressoras de Tumor/metabolismoRESUMO
The serendipitous observation of a C-H cdots, three dots, centered O hydrogen bond mediated polypeptide chain reversal in synthetic peptide helices has led to a search for the occurrence of a similar motif in protein structures. From a dataset of 634 proteins, 1304 helices terminating in a Schellman motif have been examined. The C-H triplebond O interaction between the T-4 C(alpha)H and T+1 Cz doublebond O group (C triplebond O< or =3.5A) becomes possible only when the T+1 residue adopts an extended beta conformation (T is defined as the helix terminating residue adopting an alpha(L) conformation). In all, 111 examples of this chain reversal motif have been identified and the compositional and conformational preferences at positions T-4, T, and T+1 determined. A marked preference for residues like Ser, Glu and Gln is observed at T-4 position with the motif being further stabilized by the formation of a side-chain-backbone O triplebond H-N hydrogen bond involving the side-chain of residue T-4 and the N-H group of residue T+3. In as many as 57 examples, the segment following the helix was extended with three to four successive residues in beta conformation. In a majority of these cases, the succeeding beta strand lies approximately antiparallel with the helix, suggesting that the backbone C-H triplebond O interactions may provide a means of registering helices and strands in an antiparallel orientation. Two examples were identified in which extended registry was detected with two sets of C-H cdots, three dots, centered O hydrogen bonds between (T-4) C(alpha)H triplebond O (T+1) and (T-8) C(alpha)H triplebondC doublebond O (T+3).
Assuntos
Peptídeos/química , Proteínas/química , Motivos de Aminoácidos , Carbono , Hidrogênio , Ligação de Hidrogênio , Oxigênio , Estrutura Terciária de ProteínaRESUMO
Apoptosis maintains tissue homeostasis through its ability to control cell population and has been extensively studied in human cancers. Relation of apoptosis to prognosis is still controversial. In this study, we analyzed the prognostic significance of apoptotic and mitotic indices (AI & MI) using hematoxylin and eosin stained slides by light microscopy in breast cancer patients. In our study, apoptotic index was significantly associated with predicting relapse free survival (RFS), distant recurrence free survival (DRFS) and overall survival (OS) with lesions having higher apoptotic index showing poor prognosis. Our results also point out that quantitation of apoptotic index by simple light microscopy as a routine practice along with histological diagnosis, could provide additional prognostic information in patients who are at high risk of developing recurrence with breast cancers.