Contribution of Intra- and Intermolecular Hydrogen Bonds to the Conformational Stability of Human Lysozyme^,

Abstract

In globular proteins, there are intermolecular hydrogen bonds between protein and water molecules, and between water molecules, which are bound with the proteins, in addition to intramolecular hydrogen bonds. To estimate the contribution of these hydrogen bonds to the conformational stability of a protein, the thermodynamic parameters for denaturation and the crystal structures of five Thr to Val and five Thr to Ala mutant human lysozymes were determined. The denaturation Gibbs energy (DeltaG) of Thr to Val and Thr to Ala mutant proteins was changed from 4.0 to -5.6 kJ/mol and from 1.6 to -6.3 kJ/mol, respectively, compared with that of the wild-type protein. The contribution of hydrogen bonds to the stability (DeltaDeltaG(HB)) of the Thr and other mutant human lysozymes previously reported was extracted from the observed stability changes (DeltaDeltaG) with correction for changes in hydrophobicity and side chain conformational entropy between the wild-type and mutant structures. The estimation of the DeltaDeltaG(HB) values of all mutant proteins after removal of hydrogen bonds, including protein-water hydrogen bonds, indicates a favorable contribution of the intra- and intermolecular hydrogen bonds to the protein stability. The net contribution of an intramolecular hydrogen bond (DeltaG(HB[pp])), an intermolecular one between protein and ordered water molecules (DeltaG(HB[pw])), and an intermolecular one between ordered water molecules (DeltaG(HB[ww])) could be estimated to be 8. 5, 5.2, and 5.0 kJ/mol, respectively, for a 3 A long hydrogen bond. This result shows the different contributions to protein stability of intra- and intermolecular hydrogen bonds. The entropic cost due to the introduction of a water molecule (DeltaG(H)()2(O)) could be also estimated to be about 8 kJ/mol.

Structural and Folding Dynamic Properties of the T70N Variant of Human Lysozyme

“…In the wild-type structure, that network connects Thr-52, Tyr-54, Ser-61, Asp-67, and Thr-70, and its rupture in the D67H mutant determines extensive structural modifications and pathological destabilization (3). Similar effects have been documented also for mutation at residue 70 with Ala and Val (29). The T70A substitution results in local disorder that prevents structure determination.…”

“…The analogous mutation into Val, however, induces a significant rearrangement in the region 68 -78 that preserves the local geometry and packing through the involvement of the Lys-69 side chain for the H-bonds and that of Arg-62 and Tyr-63 for interaction with the isopropyl group. Indeed, at 64.9°C and pH 2.7, mutant T70V is destabilized by 2.9 kJ/mol with respect to wild-type, whereas T70A loses 6.2 kJ/mol of stability (29). The structural elements obtained by our 1 H NMR study qualitatively account for the stability properties of T70N.…”

Structural and Dynamic Features of the MutT Protein in the Recognition of Nucleotides with the Mutagenic 8-Oxoguanine Base

“…According to previous reports, the contribution of the hydrogen bond to protein stability can be estimated as ϳ2 and 1.2 kcal/mol for hydrogen bonds between protein residues and for those between a water molecule and a protein residue, respectively (47,48). Judging from the MutT-8oxo-dGMP structure, the N119D and N119A mutants lose one hydrogen bond between the O6 of 8-oxo-dGMP and the N␦-H of Asn-119 and two hydrogen bonds involving the O6 and N7-H of 8-oxodGMP and the amide group of Asn-119, respectively (Fig.…”

“…Thus, the difference in the number of hydrogen bonds formed between MutT-G and MutT-8-oxoG complexes is only one. The contribution of one hydrogen bond to ⌬⌬G is estimated to be 2 kcal/mol (47,48).…”

Reducing the In-Vitro Electromagnetic Field Effect of Cellular Phones on Human DNA and the Intensity of Their Emitted Radiation

“…The conformation, or secondary structure of all biomolecules is intimately linked to their functional properties [17]. Conformational states are in turn controlled by hydrogen bonds which hold together the folded secondary structure of biomolecules [18]. DNA winding experiments in-vitro, like those utilized in the present study, can be considered an indirect measurement of hydrogen bond formation.…”