An automated approach for clustering an ensemble of NMR-derived protein structures into conformationally related subfamilies

Kelley, Lawrence A.; Gardner, Stephen P.; Sutcliffe, Michael J.

doi:10.1093/protein/9.11.1063

Cited by 455 publications

(371 citation statements)

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“…After the MC simulations, the dimer structures were filtered to remove structures incompatible with the dimer symmetry. Then, we clustered the remaining structures by C␣ root-meansquare (RMS) distances using NMRCLUSTER (26). Finally, the median model from the largest cluster was selected as our final predicted structure.…”

Section: Methodsmentioning

confidence: 99%

The Affinity of GXXXG Motifs in Transmembrane Helix-Helix Interactions Is Modulated by Long-range Communication

Melnyk

Kim

Curran

et al. 2004

Journal of Biological Chemistry

105

113

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Sequence motifs are responsible for ensuring the proper assembly of transmembrane (TM) helices in the lipid bilayer. To understand the mechanism by which the affinity of a common TM-TM interactive motif is controlled at the sequence level, we compared two well characterized GXXXG motif-containing homodimers, those formed by human erythrocyte protein glycophorin A (GpA, high-affinity dimer) and those formed by bacteriophage M13 major coat protein (MCP, low affinity dimer). In both constructs, the GXXXG motif is necessary for TM-TM association. Although the remaining interfacial residues (underlined) in GpA (LIXXGVXXG-VXXT) differ from those in MCP (VVXXGAXXGIXXF), molecular modeling performed here indicated that GpA and MCP dimers possess the same overall fold. Thus, we could introduce GpA interfacial residues, alone and in combination, into the MCP sequence to help decrypt the determinants of dimer affinity. Using both in vivo TOXCAT assays and SDS-PAGE gel migration rates of synthetic peptides derived from TM regions of the proteins, we found that the most distal interfacial sites, 12 residues apart (and ϳ18 Å in structural space), work in concert to control TM-TM affinity synergistically.After their biosynthesis and subsequent integration into a membrane, many transmembrane (TM) 1 helices associate with other pre-formed helices to form functional membrane protein domains (1). Specificity for a given helix-helix interaction is achieved through the appropriate presentation of complementary side chains, which serve as recognition elements between associating helices. The most highly studied, and apparently widespread, mode of association is mediated by the so-called GXXXG motif, which is known to act as a universal scaffold for the assembly of both TM helices (2-9) and soluble ␣-helices (10). The GXXXG motif, where two glycine residues are separated by any three amino acids on a helical framework, gives rise to a flat surface region on one face of the helix. This arrangement of Gly residues permits the close approach of interacting helices, whereupon extensive packing interactions take place between pairs of surrounding residues. It has been proposed that a portion of the interactive strength of GXXXGmediated associations may originate from inter-helix hydrogen bonds between C␣ hydrogens and carbonyl oxygen atoms on the adjacent helix (11).The glycophorin A transmembrane (GpA-TM) segment is a well characterized transmembrane helix dimer that associates with high affinity, principally by using a central GXXXG motif (3,12). The details of side chain-side chain packing for GpA are known in considerable detail, having been gleaned originally from extensive mutagenesis experiments (3), computer modeling (13, 14) and, subsequently, from a high-resolution structural analysis using nuclear magnetic resonance (NMR) for the GpA dimer in both detergent micelles (12) and lipid bilayers (15).Despite the high occurrence of the GXXXG motif in transmembrane helices (7), GpA-TM is the only GXXXG peptide dimer with a structure det...

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Section: Methodsmentioning

confidence: 99%

The Affinity of GXXXG Motifs in Transmembrane Helix-Helix Interactions Is Modulated by Long-range Communication

Melnyk

Kim

Curran

et al. 2004

Journal of Biological Chemistry

105

113

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“…As for loop 256-269 in Rrp43, 10 typical generated configurations were further sampled and refined using Protein Local Optimization Program (PLOP) [45]. Top 1000 ranked configurations were clustered by NMRCLUST [46] program according to the RMSD values of C-α atoms in the loop to select the representative models.…”

Section: Molecular Modeling and Structure Refinement Of Rna-free Exosmentioning

confidence: 99%

CryoEM structure of yeast cytoplasmic exosome complex

Liu

Niu

et al. 2016

Cell Res

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The eukaryotic multi-subunit RNA exosome complex plays crucial roles in 3′-to-5′ RNA processing and decay. Rrp6 and Ski7 are the major cofactors for the nuclear and cytoplasmic exosomes, respectively. In the cytoplasm, Ski7 helps the exosome to target mRNAs for degradation and turnover via a through-core pathway. However, the interaction between Ski7 and the exosome complex has remained unclear. The transaction of RNA substrates within the exosome is also elusive. In this work, we used single-particle cryo-electron microscopy to solve the structures of the Ski7-exosome complex in RNA-free and RNA-bound forms at resolutions of 4.2 Å and 5.8 Å, respectively. These structures reveal that the N-terminal domain of Ski7 adopts a structural arrangement and interacts with the exosome in a similar fashion to the C-terminal domain of nuclear Rrp6. Further structural analysis of exosomes with RNA substrates harboring 3′ overhangs of different length suggests a switch mechanism of RNA-induced exosome activation in the through-core pathway of RNA processing.

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“…The ␣IIb and ␤3 TMD dimeric structures then were filtered to remove the structures incompatible with the bilayer constraints. We then clustered the remaining structures by C␣ root mean square distances using NMR CLUSTER (25), which resulted in two equally populated clusters: one with a crossing point near the N terminus and the other with a crossing point close to C terminus. Both models were evaluated for consistency with the experimental results (see below).…”

Section: Fig 3 Isolation and Characterization Of Clonal Cell Lines mentioning

confidence: 99%

Transmembrane Domain Helix Packing Stabilizes Integrin αIIbβ3 in the Low Affinity State

Partridge

Liu²,

Kim

et al. 2005

Journal of Biological Chemistry

141

161

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Regulated changes in the affinity of integrin adhesion receptors ("activation") play an important role in numerous biological functions including hemostasis, the immune response, and cell migration. Physiological integrin activation is the result of conformational changes in the extracellular domain initiated by the binding of cytoplasmic proteins to integrin cytoplasmic domains. The conformational changes in the extracellular domain are likely caused by disruption of intersubunit interactions between the ␣ and ␤ transmembrane (TM) and cytoplasmic domains. Here, we reasoned that mutation of residues contributing to ␣/␤ interactions that stabilize the low affinity state should lead to integrin activation. Thus, we subjected the entire intracellular domain of the ␤3 integrin subunit to unbiased random mutagenesis and selected it for activated mutants. 25 unique activating mutations were identified in the TM and membrane-proximal cytoplasmic domain. In contrast, no activating mutations were identified in the more distal cytoplasmic tail, suggesting that this region is dispensable for the maintenance of the inactive state. Among the 13 novel TM domain mutations that lead to integrin activation were several informative point mutations that, in combination with computational modeling, suggested the existence of a specific TM helix-helix packing interface that maintains the low affinity state. The interactions predicted by the model were used to identify additional activating mutations in both the ␣ and ␤ TM domains. Therefore, we propose that helical packing of the ␣ and ␤ TM domains forms a clasp that regulates integrin activation.Integrin heterodimers are essential for the development and functioning of multicellular animals, because they mediate cell migration and cell adhesion and can influence gene expression and cell proliferation (1). All of the integrin heterodimers are composed of single pass Type I transmembrane (TM) 1 protein subunits ␣ and ␤. A central feature of these receptors is their capacity for rapid changes in their adhesive function mediated by changes in their ligand binding affinity, operationally defined here as "activation." The prototypical integrin, platelet ␣IIb␤3, is activated through interactions of the cytoplasmic integrin tails (ϳ20 and 47 residues for ␣ and ␤ tails, respectively) with intracellular proteins such as talin (2). These interactions initiate a long-range conformational change in the large extracellular domains (Ͼ700 residues each), resulting in high affinity binding of fibrinogen, von Willebrand factor, and fibronectin and consequently platelet aggregation and adherence to the vessel wall (1).Initial mutational studies suggested that a salt bridge between ␣IIbArg 995 and ␤3Asp 723 helps maintain the integrin in the low affinity state by forming part of an interactive face between ␣ and ␤ subunit cytoplasmic domains (3). Protein engineering studies from Springer laboratory have further advanced the idea that specific integrin ␣/␤ interactions maintain the low affinity conform...

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An automated approach for clustering an ensemble of NMR-derived protein structures into conformationally related subfamilies

Cited by 455 publications

References 0 publications

The Affinity of GXXXG Motifs in Transmembrane Helix-Helix Interactions Is Modulated by Long-range Communication

The Affinity of GXXXG Motifs in Transmembrane Helix-Helix Interactions Is Modulated by Long-range Communication

CryoEM structure of yeast cytoplasmic exosome complex

Transmembrane Domain Helix Packing Stabilizes Integrin αIIbβ3 in the Low Affinity State

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