dorsal/arxiv
View SchemaA double-deletion method to quantifying incremental binding energies in proteins from experiment. Example of a destabilizing hydrogen bonding pair
| Authors | Luis A. Campos, Santiago Cuesta-Lopez, Jon Lopez-Llano, Fernando Falo, Javier Sancho |
|---|---|
| Categories | |
| ArXiv ID | q-bio/0412017 |
| URL | https://arxiv.org/abs/q-bio/0412017 |
| DOI | 10.1529/biophysj.104.050203 |
Abstract
The contribution of a specific hydrogen bond in apoflavodoxin to protein stability is investigated by combining theory, experiment and simulation. Although hydrogen bonds are major determinants of protein structure and function, their contribution to protein stability is still unclear and widely debated. The best method so far devised to estimate the contribution of side-chain interactions to protein stability is double-mutant-cycle analysis, but the interaction energies so derived are not identical to incremental binding energies (the energies quantifying net contributions of two interacting groups to protein stability). Here we introduce double-deletion analysis of isolated residue pairs as a means to precisely quantify incremental binding. The method is exemplified by studying a surface-exposed hydrogen bond in a model protein (Asp96/Asn128 in apoflavodoxin). Combined substitution of these residues by alanines slightly destabilizes the protein, due to a decrease in hydrophobic surface burial. Subtraction of this effect, however, clearly indicates that the hydrogen-bonded groups in fact destabilize the native conformation. In addition, Molecular Dynamics simulations and classic double-mutant-cycle analysis explain quantitatively that, due to frustration, the hydrogen bond must form in the native structure because, when the two groups get approximated upon folding their binding becomes favorable. We would like to remark two facts: that this is the first time the contribution of a specific hydrogen bond to protein stability has been measured from experiment, and that more hydrogen bonds need to be analyzed in order to draw general conclusions on protein hydrogen bonds energetics. To that end, the double deletion method should be of help.
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"abstract": "The contribution of a specific hydrogen bond in apoflavodoxin to protein\nstability is investigated by combining theory, experiment and simulation.\nAlthough hydrogen bonds are major determinants of protein structure and\nfunction, their contribution to protein stability is still unclear and widely\ndebated. The best method so far devised to estimate the contribution of\nside-chain interactions to protein stability is double-mutant-cycle analysis,\nbut the interaction energies so derived are not identical to incremental\nbinding energies (the energies quantifying net contributions of two interacting\ngroups to protein stability). Here we introduce double-deletion analysis of\nisolated residue pairs as a means to precisely quantify incremental binding.\nThe method is exemplified by studying a surface-exposed hydrogen bond in a\nmodel protein (Asp96/Asn128 in apoflavodoxin). Combined substitution of these\nresidues by alanines slightly destabilizes the protein, due to a decrease in\nhydrophobic surface burial. Subtraction of this effect, however, clearly\nindicates that the hydrogen-bonded groups in fact destabilize the native\nconformation. In addition, Molecular Dynamics simulations and classic\ndouble-mutant-cycle analysis explain quantitatively that, due to frustration,\nthe hydrogen bond must form in the native structure because, when the two\ngroups get approximated upon folding their binding becomes favorable. We would\nlike to remark two facts: that this is the first time the contribution of a\nspecific hydrogen bond to protein stability has been measured from experiment,\nand that more hydrogen bonds need to be analyzed in order to draw general\nconclusions on protein hydrogen bonds energetics. To that end, the double\ndeletion method should be of help.",
"arxiv_id": "q-bio/0412017",
"authors": [
"Luis A. Campos",
"Santiago Cuesta-Lopez",
"Jon Lopez-Llano",
"Fernando Falo",
"Javier Sancho"
],
"categories": [
"q-bio.BM",
"q-bio.QM"
],
"doi": "10.1529/biophysj.104.050203",
"title": "A double-deletion method to quantifying incremental binding energies in proteins from experiment. Example of a destabilizing hydrogen bonding pair",
"url": "https://arxiv.org/abs/q-bio/0412017"
},
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