Molecular dynamics simulations of mutated Mycobacterium tuberculosis L-alanine dehydrogenase to illuminate the role of key residues.
L-Alanine dehydrogenase from Mycobacterium tuberculosis (L-MtAlaDH) catalyzes the NADH-dependent interconversion of l-alanine and pyruvate, and it is considered to be a potential target for the treatment of tuberculosis. The experiment has verified that amino acid replacement of the conserved active-site residues which have strong stability and no great changes in biological evolutionary process, such as His96 and Asp270, could lead to inactive mutants [Ågren et al., J. Mol. Biol. 377 (2008) 1161-1173]. However, the role of these conserved residues in catalytic reaction still remains unclear. Based on the crystal structures, a series of mutant structures were constructed to investigate the role of the conserved residues in enzymatic reaction by using molecular dynamics simulations. The results show that whatever the conserved residues were mutated, the protein can still convert its conformation from open state to closed state as long as NADH is present in active site. Asp270 maintains the stability of nicotinamide ring and ribose of NADH through hydrogen bond interactions, and His96 is helpful to convert the protein conformation by interactions with Gln271, whereas, they would lead to the structural rearrangement in active site and lose the catalytic activity when they were mutated. Additionally, we deduce that Met301 plays a major role in catalytic reaction due to fixing the nicotinamide ring of NADH to prevent its rotation, and we propose that Met301 would be mutated to the hydrophobic residue with large steric hindrance in side chain to test the activity of the protein in future experiment.
School of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165, China. Electronic address: baopingling@mail.qfnu.edu.cn.
Baoping
B
Ling
School of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165, China.
Siwei
S
Bi
School of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165, China.
Min
M
Sun
School of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165, China.
Zhihong
Z
Jing
School of Chemistry and Chemical Engineering, Qufu Normal University, Qufu, Shandong 273165, China.
Xiaoping
X
Li
Shandong Key Laboratory of Edible Mushroom Technology, School of Agriculture, Ludong University, Yantai, Shandong 264025, China.
Rui
R
Zhang
D050788:Alanine Dehydrogenase / Q000737:chemistry* / Q000378:metabolism; D006860:Hydrogen Bonding; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D011487:Protein Conformation; D013816:Thermodynamics
D050788:Alanine Dehydrogenase
Essential dynamics analysis; Molecular dynamics simulations; Mutated residues; Mycobacterium tuberculosis; l-Alanine dehydrogenase