scholarly journals Long-range PEG stapling: macrocyclization for increased protein conformational stability and resistance to proteolysis

2020 ◽  
Vol 1 (4) ◽  
pp. 273-280
Author(s):  
Qiang Xiao ◽  
Dallin S. Ashton ◽  
Zachary B. Jones ◽  
Katherine P. Thompson ◽  
Joshua L. Price
Keyword(s):  
Long Range ◽  
Olefin Metathesis ◽  
Quaternary Structure ◽  
Conformational Stability ◽  
Coiled Coil ◽  
Sh3 Domain ◽  
Tertiary Structures ◽  
Protein Conformational Stability

Long-range stapling of two Asn-linked PEG oligomers via olefin metathesis substantially increases the conformational stability of the WW and SH3 domain tertiary structures and the GCN4 coiled-coil quaternary structure.

Identification of a Collapsed Intermediate with Non-native Long-range Interactions on the Folding Pathway of a Pair of Fyn SH3 Domain Mutants by NMR Relaxation Dispersion Spectroscopy

2006 ◽  
Vol 363 (5) ◽  
pp. 958-976 ◽  
Author(s):  
Philipp Neudecker ◽  
Arash Zarrine-Afsar ◽  
Wing-Yiu Choy ◽  
D. Ranjith Muhandiram ◽  
Alan R. Davidson ◽  
...  
Keyword(s):  
Long Range ◽  
Nmr Relaxation ◽  
Sh3 Domain ◽  
Relaxation Dispersion ◽  
Folding Pathway ◽  
Long Range Interactions

scholarly journals Molecular basis for the fold organization and sarcomeric targeting of the muscle atrogin MuRF1

Open Biology ◽  
2014 ◽  
Vol 4 (3) ◽  
pp. 130172 ◽  
Author(s):  
Barbara Franke ◽  
Alexander Gasch ◽  
Dayté Rodriguez ◽  
Mohamed Chami ◽  
Muzamil M. Khan ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Quaternary Structure ◽  
E3 Ubiquitin Ligase ◽  
Coiled Coil ◽  
Trim Protein ◽  
Muscle Catabolism ◽  
And Function ◽  
Helical Region ◽  
Structural Significance

MuRF1 is an E3 ubiquitin ligase central to muscle catabolism. It belongs to the TRIM protein family characterized by a tripartite fold of RING, B-box and coiled-coil (CC) motifs, followed by variable C-terminal domains. The CC motif is hypothesized to be responsible for domain organization in the fold as well as for high-order assembly into functional entities. But data on CC from this family that can clarify the structural significance of this motif are scarce. We have characterized the helical region from MuRF1 and show that, contrary to expectations, its CC domain assembles unproductively, being the B2- and COS-boxes in the fold (respectively flanking the CC) that promote a native quaternary structure. In particular, the C-terminal COS-box seemingly forms an α-hairpin that packs against the CC, influencing its dimerization. This shows that a C-terminal variable domain can be tightly integrated within the conserved TRIM fold to modulate its structure and function. Furthermore, data from transfected muscle show that in MuRF1 the COS-box mediates the in vivo targeting of sarcoskeletal structures and points to the pharmacological relevance of the COS domain for treating MuRF1-mediated muscle atrophy.

Coiled-coil deformations in crystal structures: themeasles virusphosphoprotein multimerization domain as an illustrative example

2014 ◽  
Vol 70 (6) ◽  
pp. 1589-1603 ◽  
Author(s):  
David Blocquel ◽  
Johnny Habchi ◽  
Eric Durand ◽  
Marion Sevajol ◽  
François Ferron ◽  
...  
Keyword(s):  
Measles Virus ◽  
Quaternary Structure ◽  
Dynamic Equilibrium ◽  
Crystal Packing ◽  
Coiled Coil ◽  
Terminal Region ◽  
Size Exclusion ◽  
Structural Deformation ◽  
Structural Comparison ◽  
Exclusion Chromatography

The structures of two constructs of themeasles virus(MeV) phosphoprotein (P) multimerization domain (PMD) are reported and are compared with a third structure published recently by another group [Communieet al.(2013),J. Virol.87, 7166–7169]. Although the three structures all have a tetrameric and parallel coiled-coil arrangement, structural comparison unveiled considerable differences in the quaternary structure and unveiled that the three structures suffer from significant structural deformation induced by intermolecular interactions within the crystal. These results show that crystal packing can bias conclusions about function and mechanism based on analysis of a single crystal structure, and they challenge to some extent the assumption according to which coiled-coil structures can be reliably predicted from the amino-acid sequence. Structural comparison also highlighted significant differences in the extent of disorder in the C-terminal region of each monomer. The differential flexibility of the C-terminal region is also supported by size-exclusion chromatography and small-angle X-ray scattering studies, which showed that MeV PMD exists in solution as a dynamic equilibrium between two tetramers of different compaction. Finally, the possible functional implications of the flexibility of the C-terminal region of PMD are discussed.

scholarly journals Cross-linking reveals laminin coiled-coil architecture

2016 ◽  
Vol 113 (47) ◽  
pp. 13384-13389 ◽  
Author(s):  
Gad Armony ◽  
Etai Jacob ◽  
Toot Moran ◽  
Yishai Levin ◽  
Tevie Mehlman ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Self Assembly ◽  
Quaternary Structure ◽  
Coiled Coil ◽  
Surface Proteins ◽  
Cross Linking ◽  
Single Particle Reconstruction ◽  
Cross Links ◽  
Particle Reconstruction ◽  
Key Questions

Laminin, an ∼800-kDa heterotrimeric protein, is a major functional component of the extracellular matrix, contributing to tissue development and maintenance. The unique architecture of laminin is not currently amenable to determination at high resolution, as its flexible and narrow segments complicate both crystallization and single-particle reconstruction by electron microscopy. Therefore, we used cross-linking and MS, evaluated using computational methods, to address key questions regarding laminin quaternary structure. This approach was particularly well suited to the ∼750-Å coiled coil that mediates trimer assembly, and our results support revision of the subunit order typically presented in laminin schematics. Furthermore, information on the subunit register in the coiled coil and cross-links to downstream domains provide insights into the self-assembly required for interaction with other extracellular matrix and cell surface proteins.

scholarly journals Protein stability: a crystallographer's perspective

2016 ◽  
Vol 72 (2) ◽  
pp. 72-95 ◽  
Author(s):  
Marc C. Deller ◽  
Leopold Kong ◽  
Bernhard Rupp
Keyword(s):  
Protein Stability ◽  
Conformational Stability ◽  
Factors Affecting ◽  
Food Industries ◽  
Compositional Stability ◽  
Major Interest ◽  
Protein Conformational Stability ◽  
Academic Researchers ◽  
Stability Issues ◽  
Practical Level

Protein stability is a topic of major interest for the biotechnology, pharmaceutical and food industries, in addition to being a daily consideration for academic researchers studying proteins. An understanding of protein stability is essential for optimizing the expression, purification, formulation, storage and structural studies of proteins. In this review, discussion will focus on factors affecting protein stability, on a somewhat practical level, particularly from the view of a protein crystallographer. The differences between protein conformational stability and protein compositional stability will be discussed, along with a brief introduction to key methods useful for analyzing protein stability. Finally, tactics for addressing protein-stability issues during protein expression, purification and crystallization will be discussed.

scholarly journals Local water bridges and protein conformational stability

1999 ◽  
Vol 8 (10) ◽  
pp. 1982-1989 ◽  
Author(s):  
Michael Petukhov ◽  
David Cregut ◽  
Cláaaudio M. Soares ◽  
Luis Serrano
Keyword(s):  
Conformational Stability ◽  
Protein Conformational Stability ◽  
Local Water ◽  
Water Bridges

scholarly journals The SH3 domain of UNC-89 (obscurin) interacts with paramyosin, a coiled-coil protein, in Caenorhabditis elegans muscle

2016 ◽  
Vol 27 (10) ◽  
pp. 1606-1620 ◽  
Author(s):  
Hiroshi Qadota ◽  
Olga Mayans ◽  
Yohei Matsunaga ◽  
Jonathan L. McMurry ◽  
Kristy J. Wilson ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Consensus Sequence ◽  
Coiled Coil ◽  
Thick Filament ◽  
Sh3 Domain ◽  
Structural Features ◽  
Loss Of Function ◽  
Heavy Chains ◽  
Binding Partners ◽  
Two Hybrid

UNC-89 is a giant polypeptide located at the sarcomeric M-line of Caenorhabditis elegans muscle. The human homologue is obscurin. To understand how UNC-89 is localized and functions, we have been identifying its binding partners. Screening a yeast two-hybrid library revealed that UNC-89 interacts with paramyosin. Paramyosin is an invertebrate-specific coiled-coil dimer protein that is homologous to the rod portion of myosin heavy chains and resides in thick filament cores. Minimally, this interaction requires UNC-89’s SH3 domain and residues 294–376 of paramyosin and has a KD of ∼1.1 μM. In unc-89 loss-of-function mutants that lack the SH3 domain, paramyosin is found in accumulations. When the SH3 domain is overexpressed, paramyosin is mislocalized. SH3 domains usually interact with a proline-rich consensus sequence, but the region of paramyosin that interacts with UNC-89’s SH3 is α-helical and lacks prolines. Homology modeling of UNC-89’s SH3 suggests structural features that might be responsible for this interaction. The SH3-binding region of paramyosin contains a “skip residue,” which is likely to locally unwind the coiled-coil and perhaps contributes to the binding specificity.

scholarly journals Phyla-specific Correlated Dynamics in Tropomyosin

2018 ◽  
Author(s):  
J.K. James ◽  
V. Nanda
Keyword(s):  
Muscle Contraction ◽  
Dynamic Behavior ◽  
Long Range ◽  
Coiled Coil ◽  
Molecular Simulations ◽  
Actin Binding ◽  
Extensive Sequence ◽  
Dynamic Correlations ◽  
Rigid Rod ◽  
Structural Fluctuations

ABSTRACTTropomyosin (Tpm) is a continuous α-helical coiled-coil homodimer that regulates actinomyosin interactions in muscle. We examined extended molecular simulations of four Tpms, two from the vertebrate phylum Chordata (rat and pig), and two from the invertebrate Arthropoda (shrimp and lobster), and found that despite extensive sequence and structural homologyacross metazoans, dynamic behavior – particularly long range structural fluctuations – were clearly distinct between phyla. Vertebrate Tpms were flexible and sampled complex, multi-state conformational landscapes. Invertebrate Tpms were rigid, sampling highly constrained harmonic landscapes. Filtering of trajectories by PCA into essential subspaces showed significant overlap within but not between phyla. In vertebrate Tpms, hinge-regions decoupled long-range inter-helical motions and suggested distinct domains. In contrast, crustacean Tpms lacked significant long range dynamic correlations – behaving more like a single rigid rod. Although Tpm sequence and structure has highly conserved over the last 0.6-billion years since the split of ancestral bilateria into protostomes and deuterostomes, divergence seems to have occurred at the level of long-range correlated dynamics, reflecting adaptations to phyla-specific requirements of actin binding and muscle contraction.

scholarly journals Sis1 potentiates the stress response to protein aggregation and elevated temperature

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Courtney L. Klaips ◽  
Michael H. M. Gropp ◽  
Mark S. Hipp ◽  
F. Ulrich Hartl
Keyword(s):  
Stress Response ◽  
Protein Aggregation ◽  
Protein Misfolding ◽  
Mammalian Cells ◽  
Environmental Changes ◽  
Conformational Stability ◽  
General Role ◽  
Heat Stress Response ◽  
Protein Conformational Stability ◽  
Polyq Protein

AbstractCells adapt to conditions that compromise protein conformational stability by activating various stress response pathways, but the mechanisms used in sensing misfolded proteins remain unclear. Moreover, aggregates of disease proteins often fail to induce a productive stress response. Here, using a yeast model of polyQ protein aggregation, we identified Sis1, an essential Hsp40 co-chaperone of Hsp70, as a critical sensor of proteotoxic stress. At elevated levels, Sis1 prevented the formation of dense polyQ inclusions and directed soluble polyQ oligomers towards the formation of permeable condensates. Hsp70 accumulated in a liquid-like state within this polyQ meshwork, resulting in a potent activation of the HSF1 dependent stress response. Sis1, and the homologous DnaJB6 in mammalian cells, also regulated the magnitude of the cellular heat stress response, suggesting a general role in sensing protein misfolding. Sis1/DnaJB6 functions as a limiting regulator to enable a dynamic stress response and avoid hypersensitivity to environmental changes.

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