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43335-G4
Interplays of Different Interactions in Protein Structure and Function
We have performed quantum mechanical (QM) calculations and molecular dynamics (MD) and free energy (potential of mean force) simulations on certain peptides and enzymes to determine the energetics of hydrogen bonding interactions and understand the interplays of different interactions in protein structure and function.
(1) The interplay of peptide hydrogen bonds and C-H…O hydrogen bonds in stabilizing beta-sheets.
Systematical calculations have been performed for understanding the energetic roles of different hydrogen bonding interactions in stabilizing beta-sheets. The QM and ONIOM methods along with a procedure developed by us were used to estimate the contributions of individual peptide groups as well as the C-H groups to the inter-stand interactions. It was demonstrated that while the contributions of the H-bond donors are close to the H-bond energies for two interacting N-methylacetamide (NMA) molecules, the contributions of the H-bond acceptors are significantly greater. The results suggest that the interactions involving the C=O groups may provide larger stabilization effects for the beta-sheets than the N-H groups even though the C=O and N-H groups are the hydrogen bonding partners. Thus, the C=O groups in the beta-sheets may be able to fulfill much of their hydrogen bonding potential as in the case of á-helices. The origin of the highly imbalanced energetic contributions for the donor and acceptor peptide linkages was examined, and it was suggested that the inter-strand C-H• • •O=C interactions may play an important role in enhancing the energetic contributions of the H-bond acceptors.
(2) Interplay of hydrogen bonds and proton transfer in protein function: serine carboxyl peptidases
The MD and free energy simulations along with hybrid QM/MM potential were performed to study the interplay of hydrogen bonding interactions and proton transfers in the reaction catalyzed by kumamolisin-As (a serine-carboxyl peptidase) and to elucidate the catalytic mechanism and origin of substrate specificity. It was demonstrated that unlike serine peptidases that use the well-known oxyanion-hole interactions for electrostatic stabilization of the tetrahedral intermediates, the members of the sedolisin family might stabilize the tetrahedral intermediates primarily through the general acid-base mechanism. The second accomplishment of our research is the demonstration for the existence of dynamic substrate-assisted catalysis based on the computational study of kumamolisin-As. Substrate-assisted catalysis (SAC) is the process in which one or more functional groups from the substrate, in addition to those from the enzyme, may contribute to the rate acceleration for the enzyme-catalyzed reaction. Understanding the role of SAC in enzyme specificity is of considerable interest. The results of the computer simulations demonstrated that the conformational changes of distant substrate groups triggered by the bond breaking and making events of enzyme-catalyzed reactions could play an important role in the catalysis. This type of SAC involving the conformational changes of substrates was termed as dynamic SAC (to be distinguished from the standard SAC for which the conformational changes are not required). The coupling of the dynamics of substrate or protein groups to the bond breaking and making events in the promotion of the catalysis is of considerable interest and might be a general strategy in enzyme catalysis and substrate discrimination.
(3) Protein lysine methyltransferases (PKMTs): origin of product specificity
The fundamental building block of eukaryotic chromatin is the nucleosome in which 146 bp of DNA is wrapped around the core of eight histone proteins. The N-terminal tails of histone proteins are subject to a variety of post-translational covalent modifications, including histone lysine methylation catalyzed by PKMTs. These modifications form the so-called epigenetic histone code in the regulation of chromatin structure and gene expression. PKMTs may transfer one to three methyl groups from AdoMet to the å-amino group of the target lysine residue in histones. The ability of different PKMTs to direct different degrees of methylation is called product specificity (e.g., mono-, di-, or tri-methylation), and the enzymes can therefore be classified as mono-methylases, di-methylases or tri-methylases. One outstanding question for PKMTs concerns the key factors that determine the product specificity. We have performed the QM/MM MD and free energy simulations on human SET7/9 to address the question concerning the origin of the productive specificity. The experimentally determined productive specificity of SET7/9 was well reproduced by the simulations for the both wild-type and Y305F. Our results also provided additional insights into the active site interactions, including C=O…H-C hydrogen bonds, that are of importance for the enzyme's function. The mechanism for the deprotonation of the positively charged lysine residue before its methylation by the enzyme was also discussed.
