Improving Coiled-coil Stability by Optimizing Ionic Interactions

Burkhard, Peter; Ivaninskii, Sergei; Lustig, Ariel

doi:10.1016/s0022-2836(02)00114-6

Cited by 97 publications

(82 citation statements)

References 34 publications

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“…9 be consistent with work showing that the stability of a particular oligomeric state for synthetic coiled coils can be improved by consideration of the ionic interactions [24] and modulated according to the salt concentration of the buffer [25].…”

Section: Accepted M Manuscriptsupporting

confidence: 88%

Oligomerisation and thermal stability of polyvalent integrin α5β1 ligands

Kreiner

Byron

Domingues

et al. 2009

Biophysical Chemistry

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Please cite this article as: Michaela Kreiner, Olwyn Byron, Diana Domingues, Christopher F. van der Walle, Oligomerisation and thermal stability of polyvalent integrin α5β1 ligands, Biophysical Chemistry (2009Chemistry ( ), doi: 10.1016Chemistry ( /j.bpc.2009 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT2 AbstractSynthetic oligomeric integrin α5β1 ligands, specifically immobilised to surfaces, facilitate increased fibroblast cell spreading compared with that associated with the monomer. These ligands consist of a N-terminal fibronectin domain pair, a spacer and a di-, tri-or tetrameric coiled coil. However, itis not yet clear what effect fusion of the fibronectin domains has on the predicted oligomerisation of the coiled coils. Using analytical ultracentrifugation we show that the predicted tetrameric and trimeric coiled coils facilitate a corresponding ligand oligomerisation with half-dissociation at 0.7 and 0.2 µM, respectively. In contrast, the predicted dimeric coiled coil formed both dimers and trimers. Under non-reducing conditions, the unique C-terminal thiol-facilitated inter-oligomer dimerisation of the trimeric species, generating hexameric ligands. Disulphide bonding also increased helical stability during thermal unfolding. The work allows the cellular response to these clustered integrin α5β1 ligands to be more accurately interpreted, and has wider implications with respect to the utility of coiled coils as tools to facilitate protein oligomerisation.

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Section: Accepted M Manuscriptsupporting

confidence: 88%

Oligomerisation and thermal stability of polyvalent integrin α5β1 ligands

Kreiner

Byron

Domingues

et al. 2009

Biophysical Chemistry

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“…Previously elucidated factors include lateral pairing of nonpolar side chains (9-16), which is driven by stereochemical complementarity, and lateral pairing of polar side chains (9)(10)(11)(12)(13)(14)(15)(16)33), which is driven by hydrogen bonding (at least for Asn-Asn pairs). In addition, pairing preferences are influenced by the electrostatic interactions (attractive or repulsive) between ionized side chains (34)(35)(36)(37)(38)(39). Our findings indicate that vertical interactions must be considered as well if we are to understand the origins of dimerization selectivity among natural coiled-coil sequences and expand current abilities to predict and design such interactions.…”

Section: Discussionmentioning

confidence: 85%

Preferred side-chain constellations at antiparallel coiled-coil interfaces

Hadley

Testa

Woolfson

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Reliable predictive rules that relate protein sequence to structure would facilitate postgenome predictive biology and the engineering and de novo design of peptides and proteins. Through a combination of experiment and analysis of the protein data bank (PDB), we have deciphered and rationalized new rules for helixhelix interfaces of a common protein-folding and association motif, the antiparallel dimeric coiled coil. These interfaces are defined by a specific pattern of interactions among largely hydrophobic side chains often referred to as knobs-into-holes (KIH) packing: a knob from one helix inserts into a hole formed by four residues on the partner. Previous work has focused on lateral interactions within the KIH motif, for example, between an a position on one helix and a d position on the other in an antiparallel coiled coil. We show that vertical interactions within the KIH motif, such as a -a-a , are energetically important as well. The experimental and database analyses concur regarding preferred vertical combinations, which can be rationalized as leading to favorable side-chain interactions that we call constellations. The findings presented here highlight an unanticipated level of complexity in coiled-coil interactions, and our analysis of a few specific constellations illustrates a general, multipronged approach to addressing this complexity.backbone thioester exchange ͉ knobs-into-holes packing ͉ peptides ͉ protein design ͉ protein folding

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“…The highly helical nature of this peptide suggests that it is unlikely to have failed to dimerize because it was too short. Short sections of coiled coil that do not dimerize also fail to form an ␣-helix (22), whereas peptides that have an optimum sequence for formation of coiled coil only need to be 2 heptads long to dimerize (23). Other peptides that are only 4 heptads long and rich in either Lys or Glu residues form stable heterodimers when mixed but random coil monomers when kept separate (22).…”

Section: Resultsmentioning

confidence: 99%

The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

Knight

Thirumurugan

et al. 2005

Journal of Biological Chemistry

140

186

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Myosin 10 contains a region of predicted coiled coil 120 residues long. However, the highly charged nature and pattern of charges in the proximal 36 residues appear incompatible with coiled-coil formation. Circular dichroism, NMR, and analytical ultracentrifugation show that a synthesized peptide containing this region forms a stable single ␣-helix (SAH) domain in solution and does not dimerize to form a coiled coil even at millimolar concentrations. Additionally, electron microscopy of a recombinant myosin 10 containing the motor, the three calmodulin binding domains, and the fulllength predicted coiled coil showed that it was mostly monomeric at physiological protein concentration. In dimers the molecules were joined only at their extreme distal ends, and no coiled-coil tail was visible. Furthermore, the neck lengths of both monomers and dimers were much longer than expected from the number of calmodulin binding domains. In contrast, micrographs of myosin 5 heavy meromyosin obtained under the same conditions clearly showed a coiled-coil tail, and the necks were the predicted length. Thus the predicted coiled coil of myosin 10 forms a novel elongated structure in which the proximal region is a SAH domain and the distal region is a SAH domain (or has an unknown extended structure) that dimerizes only at its end. Sequence comparisons show that similar structures may exist in the predicted coiled-coil domains of myosins 6 and 7a and MyoM and could function to increase the size of the working stroke.Myosins make up a diverse superfamily of motor proteins (1). The human genome alone contains about 40 myosin genes (2). Of these, about one-third are "conventional" myosins (i.e. the well studied myosin 2), and the rest fall into about 10 different classes. The structure, properties and functions of the majority of myosin classes are poorly characterized and have largely been inferred from sequence comparisons rather than direct experiments on purified proteins (1-3).Muscle myosin 2 dimerizes through its ␣-helical coiled-coil tail. Therefore, it is commonly assumed that any myosin will also be dimeric if it contains a region predicted to be coiled coil. This assumption is dependent on the accuracy of coiled-coil prediction programs, such as COILS (4), PAIRCOIL, or MULTICOIL (5), which are also used by protein-fold prediction sites on the Web such as SMART (6).Although myosin 10 contains a region of predicted coiled coil (Fig. 1A), and is predicted to dimerize, this has not been determined experimentally. We noticed that part of the predicted coiled-coil domain of myosin 10 is highly enriched in charged residues (Fig. 1B). The proximal region consisting of 36 residues is particularly enriched in both positively and negatively charged residues, including the a and d positions of the heptad repeat (a-g) that are canonically hydrophobic residues in coiled coils (Fig. 1B). We suspected that this highly charged sequence is unlikely to form a coiled coil (7), suggesting that this part of myosin 10 may not be able to dimerize....

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Improving Coiled-coil Stability by Optimizing Ionic Interactions

Cited by 97 publications

References 34 publications

Oligomerisation and thermal stability of polyvalent integrin α5β1 ligands

Oligomerisation and thermal stability of polyvalent integrin α5β1 ligands

Preferred side-chain constellations at antiparallel coiled-coil interfaces

The Predicted Coiled-coil Domain of Myosin 10 Forms a Novel Elongated Domain That Lengthens the Head

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