Multiple steps of prion strain adaptation to a new host.

The transmission of prions across species is a critical aspect of their dissemination among mammalian hosts, including humans. This process often necessitates strain adaptation. In this study, we sought to investigate the mechanisms underlying prion adaptation while mitigating biases associated with the history of cross-species transmission of natural prion strains. To achieve this, we utilized the synthetic hamster prion strain S05. Propagation of S05 using mouse PrPC in Protein Misfolding Cyclic Amplification did not immediately overcome the species barrier. This finding underscores the involvement of factors beyond disparities in primary protein structures. Subsequently, we performed five serial passages to stabilize the incubation time to disease in mice. The levels of PrPSc increased with each passage, reaching a maximum at the third passage, and declining thereafter. This suggests that only the initial stage of adaptation is primarily driven by an acceleration in PrPSc replication. During the protracted adaptation to a new host, we observed significant alterations in the glycoform ratio and sialylation status of PrPSc N-glycans. These changes support the notion that qualitative modifications in PrPSc contribute to a more rapid disease progression. Furthermore, consistent with the decline in sialylation, a cue for "eat me" signaling, the newly adapted strain exhibited preferential colocalization with microglia. In contrast to PrPSc dynamics, the intensity of microglia activation continued to increase after the third passage in the new host. In summary, our study elucidates that the adaptation of a prion strain to a new host is a multi-step process driven by several factors.

Multiple steps of prion strain adaptation to a new host.

The transmission of prions across species is a critical aspect of their dissemination among mammalian hosts, including humans. This process often necessitates strain adaptation. In this study, we sought to investigate the mechanisms underlying prion adaptation while mitigating biases associated with the history of cross-species transmission of natural prion strains. To achieve this, we utilized the synthetic hamster prion strain S05. Propagation of S05 using mouse PrPC in Protein Misfolding Cyclic Amplification did not immediately overcome the species barrier. This finding underscores the involvement of factors beyond disparities in primary protein structures. Subsequently, we performed five serial passages to stabilize the incubation time to disease in mice. The levels of PrPSc increased with each passage, reaching a maximum at the third passage, and declining thereafter. This suggests that only the initial stage of adaptation is primarily driven by an acceleration in PrPSc replication. During the protracted adaptation to a new host, we observed significant alterations in the glycoform ratio and sialylation status of PrPSc N-glycans. These changes support the notion that qualitative modifications in PrPSc contribute to a more rapid disease progression. Furthermore, consistent with the decline in sialylation, a cue for "eat me" signaling, the newly adapted strain exhibited preferential colocalization with microglia. In contrast to PrPSc dynamics, the intensity of microglia activation continued to increase after the third passage in the new host. In summary, our study elucidates that the adaptation of a prion strain to a new host is a multi-step process driven by several factors.

Amyloid Precursor Protein Changes Arrangement in a Membrane and Its Structure Depending on the Cholesterol Content.

One of the hallmarks of Alzheimer's disease (AD) is the accumulation of amyloid beta (Aß) peptides in the brain. The processing of amyloid precursor protein (APP) into Aß is dependent on the location of APP in the membrane, membrane lipid composition and, possibly, presence of lipid rafts. In this study, we used atomic force microscopy (AFM) to investigate the interaction between transmembrane fragment APP672-726 (corresponding to Aß1-55) and its amyloidogenic mutant L723P with membranes combining liquid-ordered and liquid-disordered lipid phases. Our results demonstrated that most of the APP672-726 is located either in the liquid-disordered phase or at the boundary between ordered and disordered phases, and hardly ever in rafts. We did not notice any major changes in the domain structure induced by APP672-726. In membranes without cholesterol APP672-726, and especially its amyloidogenic mutant L723P formed annular structures and clusters rising above the membrane. Presence of cholesterol led to the appearance of concave membrane regions up to 2 nm in depth that were deeper for wild type APP672-726. Thus, membrane cholesterol regulates changes in membrane structure and permeability induced by APP that might be connected with further formation of membrane pores.

The Role of Genetic Variants in the Long Non-Coding RNA Genes MALAT1 and H19 in the Pathogenesis of Childhood Obesity.

Long non-coding RNAs (lncRNAs) play important roles in the maintenance of metabolic homeostasis. Recently, many studies have suggested that lncRNAs, such as Metastasis Associated Lung Adenocarcinoma Transcript 1 (MALAT1) and Imprinted Maternally Expressed Transcript (H19), might participate in the pathogenesis of metabolic disorders such as obesity. We conducted a case-control study with 150 Russian children and adolescents aged between 5 and 17 years old in order to assess the statistical association between the single nucleotide polymorphisms (SNPs) rs3200401 in MALAT1 and rs217727 in H19, and the risk of developing obesity in this population. We further explored the possible association of rs3200401 and rs217727 with BMI Z-score and insulin resistance. The MALAT1 rs3200401 and H19 rs217727 SNPs were genotyped using Taqman SNP genotyping assay. The MALAT1 rs3200401 SNP was identified as a risk factor for childhood obesity (p < 0.05) under the dominant and allelic models, and the CT heterozygous genotype was associated with the risk of increased BMI and with insulin resistance. The H19 rs217727 SNP had no significant association with obesity risk (all p > 0.05). Our findings thus suggest that MALAT1 SNP rs3200401 is a potential indicator of obesity susceptibility and pathogenesis in children and adolescents.

Diversity of Structural, Dynamic, and Environmental Effects Explain a Distinctive Functional Role of Transmembrane Domains in the Insulin Receptor Subfamily.

Human InsR, IGF1R, and IRR receptor tyrosine kinases (RTK) of the insulin receptor subfamily play an important role in signaling pathways for a wide range of physiological processes and are directly associated with many pathologies, including neurodegenerative diseases. The disulfide-linked dimeric structure of these receptors is unique among RTKs. Sharing high sequence and structure homology, the receptors differ dramatically in their localization, expression, and functions. In this work, using high-resolution NMR spectroscopy supported by atomistic computer modeling, conformational variability of the transmembrane domains and their interactions with surrounding lipids were found to differ significantly between representatives of the subfamily. Therefore, we suggest that the heterogeneous and highly dynamic membrane environment should be taken into account in the observed diversity of the structural/dynamic organization and mechanisms of activation of InsR, IGF1R, and IRR receptors. This membrane-mediated control of receptor signaling offers an attractive prospect for the development of new targeted therapies for diseases associated with dysfunction of insulin subfamily receptors.

Aß plaques do not protect against HSV-1 infection in a mouse model of familial Alzheimer's disease, and HSV-1 does not induce Aß pathology in a model of late onset Alzheimer's disease.

The possibility that the etiology of late onset Alzheimer's disease is linked to viral infections of the CNS has been actively debated in recent years. According to the antiviral protection hypothesis, viral pathogens trigger aggregation of Aß peptides that are produced as a defense mechanism in response to infection to entrap and neutralize pathogens. To test the causative relationship between viral infection and Aß aggregation, the current study examined whether Aß plaques protect the mouse brain against Herpes Simplex Virus 1 (HSV-1) infection introduced via a physiological route and whether HSV-1 infection triggers formation of Aß plaques in a mouse model of late-onset AD that does not develop Aß pathology spontaneously. In aged 5XFAD mice infected via eye scarification, high density of Aß aggregates did not improve survival time or rate when compared with wild type controls. In 5XFADs, viral replication sites were found in brain areas with a high density of extracellular Aß deposits, however, no association between HSV-1 and Aß aggregates could be found. To test whether HSV-1 triggers Aß aggregation in a mouse model that lacks spontaneous Aß pathology, 13-month-old hAß/APOE4/Trem2*R47H mice were infected with HSV-1 via eye scarification with the McKrae HSV-1 strain, intracranial inoculation with McKrae, intracranial inoculation after priming with LPS for 6 weeks, or intracranial inoculation with high doses of McKrae or 17syn + strains that represent different degrees of neurovirulence. No signs of Aß aggregation were found in any of the experimental groups. Instead, extensive infiltration of peripheral leukocytes was observed during the acute stage of HSV-1 infection, and phagocytic activity of myeloid cells was identified as the primary defense mechanism against HSV-1. The current results argue against a direct causative relationship between HSV-1 infection and Aß pathology.

Kinetics and Energetics of Intramolecular Electron Transfer in Single-Point Labeled TUPS-Cytochrome c Derivatives.

Electron transfer within and between proteins is a fundamental biological phenomenon, in which efficiency depends on several physical parameters. We have engineered a number of horse heart cytochrome c single-point mutants with cysteine substitutions at various positions of the protein surface. To these cysteines, as well as to several native lysine side chains, the photoinduced redox label 8-thiouredopyrene-1,3,6-trisulfonate (TUPS) was covalently attached. The long-lived, low potential triplet excited state of TUPS, generated with high quantum efficiency, serves as an electron donor to the oxidized heme c. The rates of the forward (from the label to the heme) and the reverse (from the reduced heme back to the oxidized label) electron transfer reactions were obtained from multichannel and single wavelength flash photolysis absorption kinetic experiments. The electronic coupling term and the reorganization energy for electron transfer in this system were estimated from temperature-dependent experiments and compared with calculated parameters using the crystal and the solution NMR structure of the protein. These results together with the observation of multiexponential kinetics strongly support earlier conclusions that the flexible arm connecting TUPS to the protein allows several shortcut routes for the electron involving through space jumps between the label and the protein surface.

All-d-Enantiomeric Peptide D3 Designed for Alzheimer's Disease Treatment Dynamically Interacts with Membrane-Bound Amyloid-ß Precursors.

Alzheimer's disease (AD) is a severe neurodegenerative pathology with no effective treatment known. Toxic amyloid-ß peptide (Aß) oligomers play a crucial role in AD pathogenesis. All-d-Enantiomeric peptide D3 and its derivatives were developed to disassemble and destroy cytotoxic Aß aggregates. One of the D3-like compounds is approaching phase II clinical trials; however, high-resolution details of its disease-preventing or pharmacological actions are not completely clear. We demonstrate that peptide D3 stabilizing Aß monomer dynamically interacts with the extracellular juxtamembrane region of a membrane-bound fragment of an amyloid precursor protein containing the Aß sequence. MD simulations based on NMR measurement results suggest that D3 targets the amyloidogenic region, not compromising its α-helicity and preventing intermolecular hydrogen bonding, thus creating prerequisites for inhibition of early steps of Aß conversion into ß-conformation and its toxic oligomerization. An enhanced understanding of the D3 action molecular mechanism facilitates development of effective AD treatment and prevention strategies.

Association of SNPs in Lipid Metabolism Gene Single Nucleotide Polymorphism with the Risk of Obesity in Children.

Background: Obesity is one of the most common metabolic disorders in the world, which develops due to an imbalance in energy consumption and expenditure, and both genetic and environmental factors are of great importance. We investigated the potential interactions of single nucleotide polymorphisms that might contribute to the development of polygenic obesity in children. Objective: The study involved 367 children and adolescents of both sexes aged from 4 to 18 years. The control group (normal weight) and the overweight groups included 65 and 302 children respectively. Methods: DNA for analysis was isolated from peripheral blood lymphocytes, then allelic variants rs99305069 of the FTO gene (chr16:53786615), Gln192Arg of the PON1 gene (chr7: 95308134), -250G>A of the LIPC gene (chr15: 58431740), and Ser447Ter of the LPL gene (chr8:19957678) were studied using the SNP-Express reagent kit. The results of allelic interactions were analyzed using the multifactor dimensionality reduction method. Results and Discussion: Among overweight children, the distribution of genotype and allele frequencies for the studied single nucleotide polymorphisms of the four genes corresponded to those of the control group (p > 0.05). It was found that in obese children SerSer homozygotes at the Ser447Ter polymorphism of the LPL gene, had serum triglyceride (TG) levels 2.3 times higher than in children with the same genotype from the control group. In overweight Ser447Ter heterozygotes (p < 0.0001), the TG level exceeded the control values by only 13% (p = 0.044). A two-locus genotype FTO AT/LPL SerTer, was associated with a reduced risk of childhood obesity.

Alzheimer's disease-associated ß-amyloid does not protect against herpes simplex virus 1 infection in the mouse brain.

Alzheimer's disease (AD) is a devastating fatal neurodegenerative disease. An alternative to the amyloid cascade hypothesis is that a viral infection is key to the etiology of late-onset AD, with ß-amyloid (Aß) peptides playing a protective role. In the current study, young 5XFAD mice that overexpress mutant human amyloid precursor protein with the Swedish, Florida, and London familial AD mutations were infected with one of two strains of herpes simplex virus 1 (HSV-1), 17syn+ and McKrae, at three different doses. Contrary to previous work, 5XFAD genotype failed to protect mice against HSV-1 infection. The region- and cell-specific tropisms of HSV-1 were not affected by the 5XFAD genotype, indicating that host-pathogen interactions were not altered. Seven- to ten-month-old 5XFAD animals in which extracellular Aß aggregates were abundant showed slightly better survival rate relative to their wild-type (WT) littermates, although the difference was not statistically significant. In these 5XFAD mice, HSV-1 replication centers were partially excluded from the brain areas with high densities of Aß aggregates. Aß aggregates were free of HSV-1 viral particles, and the limited viral invasion to areas with a high density of Aß aggregates was attributed to phagocytic activity of reactive microglia. In the oldest mice (12-15 months old), the survival rate did not differ between 5XFAD and WT littermates. While the current study questions the antiviral role of Aß, it neither supports nor refutes the viral etiology hypothesis of late-onset AD.

Dimeric states of transmembrane domains of insulin and IGF-1R receptors: Structures and possible role in activation.

Despite the biological significance of insulin signaling, the molecular mechanisms of activation of the insulin receptor (IR) and other proteins from its family remain elusive. Current hypothesis on signal transduction suggests ligand-triggered structural changes in the extracellular domain followed by transmembrane (TM) domains closure and dimerization leading to trans-autophosphorylation and kinase activity in intracellular segments of the receptor. Using NMR spectroscopy, we detected dimerization of isolated TM segments of IR in different membrane-mimicking environments and observed multiple signals of NH groups of protein backbone possibly corresponding to several dimer conformations. Taking available experimental data as constraints, several atomistic models of dimeric TM domains of IR and insulin-like growth factor 1 (IGF-1R) receptors were elaborated. Molecular dynamics simulations of IR ectodomain revealed noticeable collective movements potentially responsible for closure of the C-termini of FnIII-3 domains and spatial approaching of TM helices upon insulin-induced receptor activation. In addition, we demonstrated that the intracellular part of the receptor does not impose restrictions on the positioning of TM helices in the membrane. Finally, we used two independent structure prediction methods to generate a series of dimer conformations followed by their cluster analysis and dimerization free energy estimation to select the best dimer models. Biological relevance of the later was further tested via comparison of the hydrophobic organization of TM helices for both wild-type receptors and their mutants. Based on these data, the ability of several segments from other proteins to functionally replace IR and/or IGF-1R TM domains was explained.

Familial L723P Mutation Can Shift the Distribution between the Alternative APP Transmembrane Domain Cleavage Cascades by Local Unfolding of the Ε-Cleavage Site Suggesting a Straightforward Mechanism of Alzheimer's Disease Pathogenesis.

Alzheimer's disease is an age-related pathology associated with accumulation of amyloid-ß peptides, products of enzymatic cleavage of amyloid-ß precursor protein (APP) by secretases. Several familial mutations causing early onset of the disease have been identified in the APP transmembrane (TM) domain. The mutations influence production of amyloid-ß, but the molecular mechanisms of this effect are unclear. The "Australian" (L723P) mutation located in the C-termini of APP TM domain is associated with autosomal-dominant, early onset Alzheimer's disease. Herein, we describe the impact of familial L723P mutation on the structural-dynamic behavior of APP TM domain studied by high-resolution NMR in membrane-mimicking micelles and augmented by molecular dynamics simulations in explicit lipid bilayer. We found L723P mutation to cause local unfolding of the C-terminal turn of the APP TM domain helix and increase its accessibility to water required for cleavage of the protein backbone by γ-secretase in the ε-site, thus switching between alternative ("pathogenic" and "non-pathogenic") cleavage cascades. These findings suggest a straightforward mechanism of the pathogenesis associated with this mutation, and are of generic import for understanding the molecular-level events associated with APP sequential proteolysis resulting in accumulation of the pathogenic forms of amyloid-ß. Moreover, age-related onset of Alzheimer's disease can be explained by a similar mechanism, where the effect of mutation is emulated by the impact of local environmental factors, such as oxidative stress and/or membrane lipid composition. Knowledge of the mechanisms regulating generation of amyloidogenic peptides of different lengths is essential for development of novel treatment strategies of the Alzheimer's disease.

Atomistic mechanism of the constitutive activation of PDGFRA via its transmembrane domain.

Single-point mutations in the transmembrane (TM) region of receptor tyrosine kinases (RTKs) can lead to abnormal ligand-independent activation. We use a combination of computational modeling, NMR spectroscopy and cell experiments to analyze in detail the mechanism of how TM domains contribute to the activation of wild-type (WT) PDGFRA and its oncogenic V536E mutant. Using a computational framework, we scan all positions in PDGFRA TM helix for identification of potential functional mutations for the WT and the mutant and reveal the relationship between the receptor activity and TM dimerization via different interfaces. This strategy also allows us design a novel activating mutation in the WT (I537D) and a compensatory mutation in the V536E background eliminating its constitutive activity (S541G). We show both computationally and experimentally that single-point mutations in the TM region reshape the TM dimer ensemble and delineate the structural and dynamic determinants of spontaneous activation of PDGFRA via its TM domain. Our atomistic picture of the coupling between TM dimerization and PDGFRA activation corroborates the data obtained for other RTKs and provides a foundation for developing novel modulators of the pathological activity of PDGFRA.

Structural basis of the signal transduction via transmembrane domain of the human growth hormone receptor.

BACKGROUND: Prior studies of the human growth hormone receptor (GHR) revealed a distinct role of spatial rearrangements of its dimeric transmembrane domain in signal transduction across membrane. Detailed structural information obtained in the present study allowed elucidating the bases of such rearrangement and provided novel insights into receptor functioning. METHODS: We investigated the dimerization of recombinant TMD fragment GHR254-294 by means of high-resolution NMR in DPC micelles and molecular dynamics in explicit POPC membrane. RESULTS: We resolved two distinct dimeric structures of GHR TMD coexisting in membrane-mimicking micellar environment and providing left- and right-handed helix-helix association via different dimerization motifs. Based on the available mutagenesis data, the conformations correspond to the dormant and active receptor states and are distinguished by cis-trans isomerization of Phe-Pro266 bond in the transmembrane helix entry. Molecular dynamic relaxations of the structures in lipid bilayer revealed the role of the proline residue in functionally significant rearrangements of the adjacent juxtamembrane region supporting alternation between protein-protein and protein-lipid interactions of this region that can be triggered by ligand binding. Also, the importance of juxtamembrane SS bonding for signal persistency, and somewhat unusual aspects of transmembrane region interaction with water molecules were demonstrated. CONCLUSIONS: Two alternative dimeric structures of GHR TMD attributed to dormant and active receptor states interchange via allosteric rearrangements of transmembrane helices and extracellular juxtamembrane regions that support coordination between protein-protein and protein-lipid interactions. GENERAL SIGNIFICANCE: This study provides a holistic vision of GHR signal transduction across the membrane emphasizing the role of protein-lipid interactions.

The Conformation of the Epidermal Growth Factor Receptor Transmembrane Domain Dimer Dynamically Adapts to the Local Membrane Environment.

The epidermal growth factor receptor (EGFR) family is an important class of receptor tyrosine kinases, mediating a variety of cellular responses in normal biological processes and in pathological states of multicellular organisms. Different modes of dimerization of the human EGFR transmembrane domain (TMD) in different membrane mimetics recently prompted us to propose a novel signal transduction mechanism based on protein-lipid interaction. However, the experimental evidence for it was originally obtained with slightly different TMD fragments used in the two different mimetics, compromising the validity of the comparison. To eliminate ambiguity, we determined the nuclear magnetic resonance (NMR) structure of the bicelle-incorporated dimer of the EGFR TMD fragment identical to the one previously used in micelles. The NMR results augmented by molecular dynamics simulations confirm the mutual influence of the TMD and lipid environment, as is required for the proposed lipid-mediated activation mechanism. They also reveal the possible functional relevance of a subtle interplay between the concurrent processes in the lipid and protein during signal transduction.

Conformational transitions and interactions underlying the function of membrane embedded receptor protein kinases.

Among membrane receptors, the single-span receptor protein kinases occupy a broad but specific functional niche determined by distinctive features of the underlying transmembrane signaling mechanisms that are briefly overviewed on the basis of some of the most representative examples, followed by a more detailed discussion of several hierarchical levels of organization and interactions involved. All these levels, including single-molecule interactions (e.g., dimerization, liganding, chemical modifications), local processes (e.g. lipid membrane perturbations, cytoskeletal interactions), and larger scale phenomena (e.g., effects of membrane surface shape or electrochemical potential gradients) appear to be closely integrated to achieve the observed diversity of the receptor functioning. Different species of receptor protein kinases meet their specific functional demands through different structural features defining their responses to stimulation, but certain common patterns exist. Signaling by receptor protein kinases is typically associated with the receptor dimerization and clustering, ligand-induced rearrangements of receptor domains through allosteric conformational transitions with involvement of lipids, release of the sequestered lipids, restriction of receptor diffusion, cytoskeleton and membrane shape remodeling. Understanding of complexity and continuity of the signaling processes can help identifying currently neglected opportunities for influencing the receptor signaling with potential therapeutic implications. This article is part of a Special Issue entitled: Interactions between membrane receptors in cellular membranes edited by Kalina Hristova.

New Insights into Molecular Organization of Human Neuraminidase-1: Transmembrane Topology and Dimerization Ability.

Neuraminidase 1 (NEU1) is a lysosomal sialidase catalyzing the removal of terminal sialic acids from sialyloconjugates. A plasma membrane-bound NEU1 modulating a plethora of receptors by desialylation, has been consistently documented from the last ten years. Despite a growing interest of the scientific community to NEU1, its membrane organization is not understood and current structural and biochemical data cannot account for such membrane localization. By combining molecular biology and biochemical analyses with structural biophysics and computational approaches, we identified here two regions in human NEU1 - segments 139-159 (TM1) and 316-333 (TM2) - as potential transmembrane (TM) domains. In membrane mimicking environments, the corresponding peptides form stable α-helices and TM2 is suited for self-association. This was confirmed with full-size NEU1 by co-immunoprecipitations from membrane preparations and split-ubiquitin yeast two hybrids. The TM2 region was shown to be critical for dimerization since introduction of point mutations within TM2 leads to disruption of NEU1 dimerization and decrease of sialidase activity in membrane. In conclusion, these results bring new insights in the molecular organization of membrane-bound NEU1 and demonstrate, for the first time, the presence of two potential TM domains that may anchor NEU1 in the membrane, control its dimerization and sialidase activity.

Cell-free expression of the APP transmembrane fragments with Alzheimer's disease mutations using algal amino acid mixture for structural NMR studies.

Structural investigations need ready supply of the isotope labeled proteins with inserted mutations n the quantities sufficient for the heteronuclear NMR. Though cell-free expression system has been widely used in the past years, high startup cost and complex compound composition prevent many researches from the developing this technique, especially for membrane protein production. Here we demonstrate the utility of a robust, cost-optimized cell-free expression technique for production of the physiologically important transmembrane fragment of amyloid precursor protein, APP686-726, containing Alzheimer's disease mutations in the juxtamembrane (E693G, Arctic form) and the transmembrane parts (V717G, London form, or L723P, Australian form). The protein cost was optimized by varying the FM/RM ratio as well as the amino acid concentration. We obtained the wild-type and mutant transmembrane fragments in the pellet mode of continuous exchange cell-free system consuming only commercial algal mixture of the (13)C,(15)N-labeled amino acids. Scaling up analytical tests, we achieved milligram quantity yields of isotope labeled wild-type and mutant APP686-726 for structural studies by high resolution NMR spectroscopy in membrane mimicking environment. The described approach has from 5 to 23-fold cost advantage over the bacterial expression methods described earlier and 1.5 times exceeds our previous result obtained with the longer APP671-726WT fragment.

Alternative packing of EGFR transmembrane domain suggests that protein-lipid interactions underlie signal conduction across membrane.

The human epidermal growth factor receptor (EGFR) of HER/ErbB receptor tyrosine kinase family mediates a broad spectrum of cellular responses transducing biochemical signals via lateral dimerization in plasma membrane, while inactive receptors can exist in both monomeric and dimeric forms. Recently, the dimeric conformation of the helical single-span transmembrane domains of HER/ErbB employing the relatively polar N-terminal motifs in a fashion permitting proper kinase activation was experimentally determined. Here we describe the EGFR transmembrane domain dimerization via an alternative weakly polar C-terminal motif A(661)xxxG(665) presumably corresponding to the inactive receptor state. During association, the EGFR transmembrane helices undergo a structural adjustment with adaptation of inter-molecular polar and hydrophobic interactions depending upon the surrounding membrane properties that directly affect the transmembrane helix packing. This might imply that signal transduction through membrane and allosteric regulation are inclusively mediated by coupled protein-protein and protein-lipid interactions, elucidating paradoxically loose linkage between ligand binding and kinase activation.

HER2 Transmembrane Domain Dimerization Coupled with Self-Association of Membrane-Embedded Cytoplasmic Juxtamembrane Regions.

Receptor tyrosine kinases of the human epidermal growth factor receptor (HER or ErbB) family transduce biochemical signals across plasma membrane, playing a significant role in vital cellular processes and in various cancers. Inactive HER/ErbB receptors exist in equilibrium between the monomeric and unspecified pre-dimerized states. After ligand binding, the receptors are involved in strong lateral dimerization with proper assembly of their extracellular ligand-binding, single-span transmembrane, and cytoplasmic kinase domains. The dimeric conformation of the HER2 transmembrane domain that is believed to support the cytoplasmic kinase domain configuration corresponding to the receptor active state was previously described in lipid bicelles. Here we used high-resolution NMR spectroscopy in another membrane-mimicking micellar environment and identified an alternative HER2 transmembrane domain dimerization coupled with self-association of membrane-embedded cytoplasmic juxtamembrane region. Such a dimerization mode appears to be capable of effectively inhibiting the receptor kinase activity. This finding refines the molecular mechanism regarding the signal propagation steps from the extracellular to cytoplasmic domains of HER/ErbB receptors.