Context Dependence of Mutational Effects in a Protein: The Crystal Structures of the V35I, I47V and V35I/I47V Gene V Protein Core Mutants

Zhang, Hong; Skinner, Matthew M.; Sandberg, Warren S.; Wang, Andrew H.‐J.; Terwilliger, Thomas C.

doi:10.1006/jmbi.1996.0309

Cited by 19 publications

(12 citation statements)

References 40 publications

(74 reference statements)

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“…They also showed that the effect of each of mutations I52L and L105V depended on the side-chain in the other position. Such dependence on the structural context for mutations in the core of proteins has already been described (Zhang et al, 1996). The unfolding of the synthetases was monitored by their mean partition coef®cient K av in experiments of fast-SEC, as described in the legend to Figure 5.…”

Section: Coupling Between Mutations I52l and L105v And Context Effectmentioning

confidence: 79%

Experimental evolution of a dense cluster of residues in tyrosyl-tRNA synthetase: quantitative effects on activity, stability and dimerization 1 1Edited by A. R. Fersht

Park

Guez

Bedouelle

1999

Journal of Molecular Biology

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A dense cluster of eight residues was identi®ed at the crossing of two a-helices in tyrosyl-tRNA synthetase (TyrRS) from the thermophile Bacillus stearothermophilus. Its mechanism of evolution was characterized. Four residues of this cluster are not conserved in TyrRS from the mesophile Escherichia coli. The corresponding mutations were constructed in TyrRS(Á1), a derivative of TyrRS from B. stearothermophilus in which the anticodon binding domain is deleted. Mutations I52L (i.e. Ile52 into Leu), M55L and L105V did not affect the activity of TyrRS(Á1) in the pyrophosphate exchange reaction whereas T51P increased it. The kinetic stabilities of TyrRS(Á1) and its mutant derivatives at 68.5 C were determined from experiments of irreversible thermal precipitation. They were in the order L105V < I52L < T51P < Wild Type 4 M55L; mutation I52L partially compensated L105V in these experiments whereas M55L was coupled neither to I52L nor to L105V. Mutations I52L and L105V affected the stability of the dimeric TyrRS(Á1) at different steps of its unfolding by urea, monitored under equilibrium conditions by spectro¯uorometry or size exclusion chromatography. I52L destabilized the association between the subunits even though residue Ile52 is more than 20 A Ê away from the subunit interface. L105V destabilized the monomeric intermediate of unfolding. The two mutational pathways, going from the wildtype TyrRS(Á1) to the I52L-L105V double mutant through each of the single mutants were not equivalent for the stability of the monomeric intermediate and for the total stability of the dimer. One pathway contained two neutral steps whereas the other pathway contained a destabilizing step followed by a stabilizing step. Mutation I52L allowed L105V along the ®rst pathway and compensated it along the second pathway. Thus, the effects of I52L and L105V on stability depended on the structural context. The gain in activity due to T51P was at the expense of a slight destabilization.

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Section: Coupling Between Mutations I52l and L105v And Context Effectmentioning

confidence: 79%

Experimental evolution of a dense cluster of residues in tyrosyl-tRNA synthetase: quantitative effects on activity, stability and dimerization 1 1Edited by A. R. Fersht

Park

Guez

Bedouelle

1999

Journal of Molecular Biology

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“…The election of the properties was inspired by the results of different site-directed mutagenesis studies, [11][12][13][14]21 Figure 3. Dependence of the probability of compensatory mutations on the physico-chemical properties of the compensated mutations.…”

Section: Discussionmentioning

confidence: 99%

“…For example, mutation of residues involved in complex interaction networks, like protein core residues, are particularly difficult to compensate. 11,13,14 We have characterised CPDs in terms of (i) structure context, and (ii) intrinsic nature of the amino acid change. We have then compared the resulting property distributions with those from a set of disease-associated mutations, to which we will refer as pathogenic deviations (PDs).…”

Section: Introductionmentioning

confidence: 99%

“…This is a natural approach, as recent studies show that about 80% of pathological mutations result in protein stability changes, 9 while only an approximate 10% will interfere with the formation of normal quaternary interactions. 10 Our approach is inspired by site-directed mutagenesis experiments [11][12][13][14] that treat, within a thermodynamic framework, the problem of compensating damaging mutations. In these studies it is shown that certain characteristics of the latter are critical when considering the rescuing ability of compensatory mutations.…”

Section: Introductionmentioning

confidence: 99%

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Characterization of Compensated Mutations in Terms of Structural and Physico-Chemical Properties

Ferrer-Costa

Orozco

Cruz

2007

Journal of Molecular Biology

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“…However, other surface amino acids not implicated directly in DNA binding seem to have effects on the property of the GVP or the viability of the phage. Although the relation between the mutation of interior hydrophobic amino acids and protein stability has been studied extensively (Skinner & Terwilliger, 1996;Zhang et al, 1996), it is less clear how surface mutations may affect the protein stability. In this paper, we analyzed three such surface mutants, L32R, K69H, and R82C.…”

Section: Discussionmentioning

confidence: 99%

Analyses of the stability and function of three surface mutants (R82C, K69H, and L32R) of the gene V protein from Ff phage by X‐ray crystallography

Gao

Zhang

et al. 1997

Protein Science

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The high-resolution crystal structure of the gene V protein (GVP) from the Ff filamentous phages (M13, fl, fd) has been solved recently for the wild-type and two surface mutant (Y41F and Y41H) proteins, leading to a plausible model for the polymeric GVP-ssDNA complex (Guan Y, Zhang H, Wang AHJ, 1995, Protein Sci 4187-197). The model of the complex shows extensive contacts between neighboring dimer GVPs involving electrostatic interactions between the K69 from one and the D79 and R82 from the next dimer. In addition, hydrophobic interactions between the amino acids L32 and L44 from one and G23 from the next dimer also contribute to the dimer-dimer interactions. Mutations at the L32, K69, and R82 amino acid sites generally destabilize the protein and many of these affect the function of the phage. We have studied the structural effects of three mutant proteins involving those sites, i.e., L32R, K69H, and R82C, by X-ray crystallographic analysis at 2.0 8, resolution. In L32R GVP, the structural perturbation is localized, whereas in K69H and R82C GVPs, some long-range effects are also detected in addition to the local perturbation. We have interpreted the protein stability and the functional properties associated with those mutations in terms of the observed structural perturbations.

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Context Dependence of Mutational Effects in a Protein: The Crystal Structures of the V35I, I47V and V35I/I47V Gene V Protein Core Mutants

Cited by 19 publications

References 40 publications

Experimental evolution of a dense cluster of residues in tyrosyl-tRNA synthetase: quantitative effects on activity, stability and dimerization 1 1Edited by A. R. Fersht

Experimental evolution of a dense cluster of residues in tyrosyl-tRNA synthetase: quantitative effects on activity, stability and dimerization 1 1Edited by A. R. Fersht

Characterization of Compensated Mutations in Terms of Structural and Physico-Chemical Properties

Analyses of the stability and function of three surface mutants (R82C, K69H, and L32R) of the gene V protein from Ff phage by X‐ray crystallography

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