Positive Contribution of Hydration Structure on the Surface of Human Lysozyme to the Conformational Stability

Jun Funahashi,Kazufumi Takano,Yuriko Yamagata,Katsuhide Yutani

doi:10.1074/jbc.m110728200

Jun Funahashi, Kazufumi Takano + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m110728200

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Abstract

Water molecules make a hydration structure with the network of hydrogen bonds, covering on the surface of proteins. To quantitatively estimate the contribution of the hydration structure to protein stability, a series of hydrophilic mutant human lysozymes (Val to Ser, Tyr, Asp, Asn, and Arg) modified at three different positions on the surface, which are located in the alpha-helix (Val-110), the beta-sheet (Val-2), and the loop (Val-74), were constructed. Their thermodynamic parameters of denaturation and crystal structures were examined by calorimetry and by x-ray crystallography at 100 K, respectively. The introduced polar residues made hydrogen bonds with protein atoms and/or water molecules, sometimes changing the hydration structure around the mutation site. Changes in the stability of the mutant proteins can be evaluated by a unique equation that considers the conformational changes resulting from the substitutions. Using this analysis, the relationship between the changes in the stabilities and the hydration structures for mutant human lysozymes substituted on the surface could be quantitatively estimated. The analysis indicated that the hydration structure on protein surface plays an important role in determining the conformational stability of the protein.

Highlights

Water is the natural medium for protein molecules and has a significant influence on the dynamics, stability, and function of the molecules [1, 2]
Stability of Mutant Human Lysozymes—The hydrophilic mutant proteins substituted at position 2 (V2S, V2Y, V2D, V2N, V2R), position 74 (V74S, V74Y, V74D, V74N, V74R), and position 110 (V110Y, V110D, V110N, V110R) were examined
It has been proposed that the changes in stability of each mutant human lysozyme are represented by a unique equation, considering the conformational changes caused by the mutations [18, 19]

Summary

Introduction

Water is the natural medium for protein molecules and has a significant influence on the dynamics, stability, and function of the molecules [1, 2]. As the first step in understanding the contribution of the hydration structure to protein stability, the relationship between the changes in stability and hydration structure resulting from the amino acid substitution on the protein surface, measured by physicochemical experiments and cryogenic x-ray analysis, respectively, should be elucidated. The contributions of even the same kind of substitutions on the surface of proteins to their stabilities have been changed depending on the environment of the mutation sites [14] Considering these facts, systematic surveys are necessary to understand the role of the hydration structure. The contributions of several stabilization factors to the stabilities of mutant human lysozymes have been evaluated by a unique equation considering the conformational changes caused by the substitutions (18 –20). The role of surface hydrophilic residues and hydration structures in the conformational stability of a protein will be discussed along with the changes in the stability and structure caused by the substitution

Results

Discussion

Conclusion