Ranking docked models of protein-protein complexes using predicted partner-specific protein-protein interfaces

Cells and tissues are continuously exposed to both chemical and physical stimuli and dynamically adapt and respond to this variety of external cues to ensure cellular homeostasis, regulated development and tissue-specific differentiation. Alterations of these pathways promote disease progression—a prominent example being cancer. Rho GTPases are key regulators of the remodeling of cytoskeleton and cell membranes and their coordination and integration with different biological processes, including cell polarization and motility, as well as other signaling networks such as growth signaling and proliferation. Apart from the control of GTP–GDP cycling, Rho GTPase activity is spatially and temporally regulated by post-translation modifications (PTMs) and their assembly onto specific protein complexes, which determine their controlled activity at distinct cellular compartments. Although Rho GTPases were traditionally conceived as targeted from the cytosol to the plasma membrane to exert their activity, recent research demonstrates that active pools of different Rho GTPases also localize to endomembranes and the nucleus. In this review, we discuss how PTM-driven modulation of Rho GTPases provides a versatile mechanism for their compartmentalization and functional regulation. Understanding how the subcellular sorting of active small GTPase pools occurs and what its functional significance is could reveal novel therapeutic opportunities.

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Metallocluster transactions: dynamic protein interactions guide the biosynthesis of Fe–S clusters in bacteria

Biochemical Society Transactions ◽

10.1042/bst20180365 ◽

2018 ◽

Vol 46 (6) ◽

pp. 1593-1603 ◽

Cited By ~ 9

Author(s):

Chenkang Zheng ◽

Patricia C. Dos Santos

Keyword(s):

Protein Interactions ◽

Protein Complexes ◽

Specific Protein ◽

Biological Synthesis ◽

Cluster Assembly ◽

Sequence Motifs ◽

Protein Protein Interactions ◽

Cysteine Desulfurase ◽

Wide Range ◽

Domains Of Life

Iron–sulfur (Fe–S) clusters are ubiquitous cofactors present in all domains of life. The chemistries catalyzed by these inorganic cofactors are diverse and their associated enzymes are involved in many cellular processes. Despite the wide range of structures reported for Fe–S clusters inserted into proteins, the biological synthesis of all Fe–S clusters starts with the assembly of simple units of 2Fe–2S and 4Fe–4S clusters. Several systems have been associated with the formation of Fe–S clusters in bacteria with varying phylogenetic origins and number of biosynthetic and regulatory components. All systems, however, construct Fe–S clusters through a similar biosynthetic scheme involving three main steps: (1) sulfur activation by a cysteine desulfurase, (2) cluster assembly by a scaffold protein, and (3) guided delivery of Fe–S units to either final acceptors or biosynthetic enzymes involved in the formation of complex metalloclusters. Another unifying feature on the biological formation of Fe–S clusters in bacteria is that these systems are tightly regulated by a network of protein interactions. Thus, the formation of transient protein complexes among biosynthetic components allows for the direct transfer of reactive sulfur and Fe–S intermediates preventing oxygen damage and reactions with non-physiological targets. Recent studies revealed the importance of reciprocal signature sequence motifs that enable specific protein–protein interactions and consequently guide the transactions between physiological donors and acceptors. Such findings provide insights into strategies used by bacteria to regulate the flow of reactive intermediates and provide protein barcodes to uncover yet-unidentified cellular components involved in Fe–S metabolism.

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Atomic-level characterization of protein–protein association

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1815431116 ◽

2019 ◽

Vol 116 (10) ◽

pp. 4244-4249 ◽

Cited By ~ 59

Author(s):

Albert C. Pan ◽

Daniel Jacobson ◽

Konstantin Yatsenko ◽

Duluxan Sritharan ◽

Thomas M. Weinreich ◽

...

Keyword(s):

Structural Information ◽

Protein Complexes ◽

Md Simulations ◽

Atomic Level ◽

Conformational Ensemble ◽

Native State ◽

Protein Interfaces ◽

Study Association ◽

Functionally Diverse

Despite the biological importance of protein–protein complexes, determining their structures and association mechanisms remains an outstanding challenge. Here, we report the results of atomic-level simulations in which we observed five protein–protein pairs repeatedly associate to, and dissociate from, their experimentally determined native complexes using a molecular dynamics (MD)–based sampling approach that does not make use of any prior structural information about the complexes. To study association mechanisms, we performed additional, conventional MD simulations, in which we observed numerous spontaneous association events. A shared feature of native association for these five structurally and functionally diverse protein systems was that if the proteins made contact far from the native interface, the native state was reached by dissociation and eventual reassociation near the native interface, rather than by extensive interfacial exploration while the proteins remained in contact. At the transition state (the conformational ensemble from which association to the native complex and dissociation are equally likely), the protein–protein interfaces were still highly hydrated, and no more than 20% of native contacts had formed.

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Improved strategy for isoleucine 1H/13C methyl labeling in Pichia pastoris

Journal of Biomolecular NMR ◽

10.1007/s10858-019-00281-1 ◽

2019 ◽

Vol 73 (12) ◽

pp. 687-697 ◽

Cited By ~ 1

Author(s):

Rustam Ali ◽

Lindsay D. Clark ◽

Jacob A. Zahm ◽

Andrew Lemoff ◽

Karthik Ramesh ◽

...

Keyword(s):

Pichia Pastoris ◽

Isotope Labeling ◽

Protein Complexes ◽

Cost Effective ◽

Specific Protein ◽

Site Specific ◽

Methyl Labeling ◽

Structure And Dynamics ◽

Eukaryotic Proteins ◽

The Cost

Abstract Site specific methyl labeling combined with methyl TROSY offers a powerful NMR approach to study structure and dynamics of proteins and protein complexes of high molecular weight. Robust and cost-effective methods have been developed for site specific protein 1H/13C methyl labeling in an otherwise deuterated background in bacteria. However, bacterial systems are not suitable for expression and isotope labeling of many eukaryotic and membrane proteins. The yeast Pichia pastoris (P. pastoris) is a commonly used host for expression of eukaryotic proteins, and site-specific methyl labeling of perdeuterated eukaryotic proteins has recently been achieved with this system. However, the practical utility of methyl labeling and deuteration in P. pastoris is limited by high costs. Here, we describe an improved method for 1H/13C-labeling of the δ-methyl group of isoleucine residues in a perdeuterated background, which reduces the cost by ≥ 50% without compromising the efficiency of isotope enrichment. We have successfully implemented this method to label actin and a G-protein coupled receptor. Our approach will facilitate studies of the structure and dynamics of eukaryotic proteins by NMR spectroscopy.

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Hic-5, an adaptor-like nuclear receptor coactivator

Nuclear Receptor Signaling ◽

10.1621/nrs.04019 ◽

2006 ◽

Vol 4 (1) ◽

pp. nrs.04019 ◽

Cited By ~ 10

Author(s):

Marjet D. Heitzer ◽

Donald B. DeFranco

Keyword(s):

Nuclear Receptor ◽

Protein Complexes ◽

Specific Protein ◽

Nuclear Receptor Coactivator ◽

Group Iii ◽

Protein Protein Interaction ◽

Direct Modification ◽

Nuclear Receptor Coactivators ◽

Multiple Protein ◽

Receptor Interacting Proteins

In recent years, numerous nuclear receptor-interacting proteins have been identified that influence nuclear transcription through their direct modification of chromatin. Along with coactivators that possess histone acetyltransferase (HAT) or methyltransferase activity, other coactivators that lack recognizable chromatin-modifying activity have been discovered whose mechanism of action is largely unknown. The presence of multiple protein-protein interaction motifs within mechanistically undefined coactivators suggests that they function as adaptor molecules, either recruiting or stabilizing promoter-specific protein complexes. This perspective will focus on a family of nuclear receptor coactivators (i.e., group III LIM domain proteins related to paxillin) that appear to provide a scaffold to stabilize receptor interactions with chromatin-modifying coregulators.

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The Maturation Pathway of Nickel Urease

Inorganics ◽

10.3390/inorganics7070085 ◽

2019 ◽

Vol 7 (7) ◽

pp. 85 ◽

Cited By ~ 3

Author(s):

Yap Shing Nim ◽

Kam-Bo Wong

Keyword(s):

Protein Interactions ◽

Protein Complexes ◽

Toxic Metal ◽

Specific Protein ◽

Accessory Proteins ◽

Protein Protein Interactions ◽

Nickel Ions ◽

Hydrogenase Maturation ◽

Maturation Factor

Maturation of urease involves post-translational insertion of nickel ions to form an active site with a carbamylated lysine ligand and is assisted by urease accessory proteins UreD, UreE, UreF and UreG. Here, we review our current understandings on how these urease accessory proteins facilitate the urease maturation. The urease maturation pathway involves the transfer of Ni2+ from UreE → UreG → UreF/UreD → urease. To avoid the release of the toxic metal to the cytoplasm, Ni2+ is transferred from one urease accessory protein to another through specific protein–protein interactions. One central theme depicts the role of guanosine triphosphate (GTP) binding/hydrolysis in regulating the binding/release of nickel ions and the formation of the protein complexes. The urease and [NiFe]-hydrogenase maturation pathways cross-talk with each other as UreE receives Ni2+ from hydrogenase maturation factor HypA. Finally, the druggability of the urease maturation pathway is reviewed.

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Development of Sindbis Viruses Encoding nsP2/GFP Chimeric Proteins and Their Application for Studying nsP2 Functioning

Journal of Virology ◽

10.1128/jvi.02746-06 ◽

2007 ◽

Vol 81 (10) ◽

pp. 5046-5057 ◽

Cited By ~ 50

Author(s):

Svetlana Atasheva ◽

Rodion Gorchakov ◽

Robert English ◽

Ilya Frolov ◽

Elena Frolova

Keyword(s):

Virus Replication ◽

Sindbis Virus ◽

Fluorescent Protein ◽

Protein Complexes ◽

Chimeric Protein ◽

Specific Protein ◽

Heterologous Proteins ◽

Peptide Tags ◽

Infected Cells ◽

Green Fluorescent

ABSTRACT Sindbis virus (SINV) is one of almost 30 currently known alphaviruses. In infected cells, it produces only a few proteins that function in virus replication and interfere with the development of the antiviral response. One of the viral nonstructural proteins, nsP2, not only exhibits protease and RNA helicase activities that are directly involved in viral RNA replication but also plays critical roles in the development of transcriptional and translational shutoffs in the SINV-infected cells. These multiple activities of nsP2 complicate investigations of this protein's functions and further understanding of its structure. Using a transposon-based approach, we generated a cDNA library of SINV genomes with a green fluorescent protein (GFP) gene randomly inserted into nsP2 and identified a number of sites that can be used for GFP cloning without a strong effect on virus replication. Recombinant SIN viruses encoding nsP2/GFP chimeric protein were capable of growth in tissue culture and interfering with cellular functions. SINV, expressing GFP in the nsP2, was used to isolate nsP2-specific protein complexes formed in the cytoplasm of the infected cells. These complexes contained viral nsPs, all of the cellular proteins that we previously coisolated with SINV nsP3, and some additional protein factors that were not found before in detectable concentrations. The random insertion library-based approach, followed by the selection of the viable variants expressing heterologous proteins, can be applied for mapping the domain structure of the viral nonstructural and structural proteins, cloning of peptide tags for isolation of the protein-specific complexes, and studying their formation by using live-cell imaging. This approach may also be applicable to presentation of additional antigens and retargeting of viruses to new receptors.

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Negative autoregulation of the neu gene is mediated by a novel enhancer.

Molecular and Cellular Biology ◽

10.1128/mcb.12.6.2739 ◽

1992 ◽

Vol 12 (6) ◽

pp. 2739-2748 ◽

Cited By ~ 20

Author(s):

X Y Zhao ◽

M C Hung

Keyword(s):

Protein Complexes ◽

Feedback Regulation ◽

Specific Protein ◽

Initiation Codon ◽

Enhancer Activity ◽

Enhancer Element ◽

Nuclear Extracts ◽

Gel Shift ◽

Neu Oncogene ◽

Transformed Cell Lines

In an attempt to study potential feedback regulation of the neu oncogene, we have found that the neu oncogene product specifically represses its own promoter activity. Deletion analysis indicated a 140-bp region (nucleotides -312 to -173 relative to the ATG initiation codon) in the rat neu promoter responsible for neu autorepression. Gel shift assays and methylation interference analysis further demonstrated that a GGTGGGGGGG sequence (nucleotides -243 to -234 relative to the ATG initiation codon) in this 140-bp region interacts with specific protein complexes. The GGTGGGGGGG sequence (GTG element), which functions as an enhancer, is sufficient to cause neu-mediated repression in a heterologous promoter. Furthermore, it produces different gel shift patterns with nuclear extracts from neu-transformed cell lines and their parental lines, suggesting that a transcriptional factor(s) interacting with this enhancer element has been perturbed by the introduction of neu. Taken together, the data presented in this report show that (i) the neu oncogene product autorepresses its own promoter, (ii) the neu promoter contains a novel enhancer, and (iii) neu autorepression is mediated through this enhancer, likely by inhibition of the enhancer activity.

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Hypervariable Domains of nsP3 Proteins of New World and Old World Alphaviruses Mediate Formation of Distinct, Virus-Specific Protein Complexes

Journal of Virology ◽

10.1128/jvi.02853-12 ◽

2012 ◽

Vol 87 (4) ◽

pp. 1997-2010 ◽

Cited By ~ 37

Author(s):

N. J. Foy ◽

M. Akhrymuk ◽

I. Akhrymuk ◽

S. Atasheva ◽

A. Bopda-Waffo ◽

...

Keyword(s):

New World ◽

Protein Complexes ◽

Specific Protein ◽

Old World ◽

Distinct Virus

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Conservation of coevolving protein interfaces bridges prokaryote–eukaryote homologies in the twilight zone

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1611861114 ◽

2016 ◽

Vol 113 (52) ◽

pp. 15018-15023 ◽

Cited By ~ 24

Author(s):

Juan Rodriguez-Rivas ◽

Simone Marsili ◽

David Juan ◽

Alfonso Valencia

Keyword(s):

Protein Interactions ◽

Protein Complexes ◽

Accurate Information ◽

Twilight Zone ◽

Sequence Information ◽

Protein Protein Interactions ◽

Sequence Alignments ◽

Multiple Sequence ◽

Protein Interfaces ◽

Recent Developments

Protein–protein interactions are fundamental for the proper functioning of the cell. As a result, protein interaction surfaces are subject to strong evolutionary constraints. Recent developments have shown that residue coevolution provides accurate predictions of heterodimeric protein interfaces from sequence information. So far these approaches have been limited to the analysis of families of prokaryotic complexes for which large multiple sequence alignments of homologous sequences can be compiled. We explore the hypothesis that coevolution points to structurally conserved contacts at protein–protein interfaces, which can be reliably projected to homologous complexes with distantly related sequences. We introduce a domain-centered protocol to study the interplay between residue coevolution and structural conservation of protein–protein interfaces. We show that sequence-based coevolutionary analysis systematically identifies residue contacts at prokaryotic interfaces that are structurally conserved at the interface of their eukaryotic counterparts. In turn, this allows the prediction of conserved contacts at eukaryotic protein–protein interfaces with high confidence using solely mutational patterns extracted from prokaryotic genomes. Even in the context of high divergence in sequence (the twilight zone), where standard homology modeling of protein complexes is unreliable, our approach provides sequence-based accurate information about specific details of protein interactions at the residue level. Selected examples of the application of prokaryotic coevolutionary analysis to the prediction of eukaryotic interfaces further illustrate the potential of this approach.

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