Evidence that the amino acid residue Cys117 of chloroplastic monodehydroascorbate reductase is involved in its activity and structural stability.
Monodehydroascorbate reductase (MDAR; EC 1.6.5.4) is crucial for AsA regeneration and essential for maintaining the reduced pool of AsA. And the amino acid residue C117 of chloroplastic MDAR is the conserved cysteine residue in MDAR isoforms. A series mutation of conserved amino acid residue cysteine117 (C117) was constructed to investigate its role in MDAR structural stability and activity. Our study revealed that mutation in this conserved residue could cause pronounced loss of activity and conformational changes. Spectroscopic experiments indicated that these mutations influenced transition from the molten globule intermediate to the native state in folding process. These results suggested that amino acid residue C117 played a relatively important role in keeping MDAR structural stability and activity.
Li
Feng
F
College of Life Science, State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai'an, Shandong 271018, PR China.
Wu
Qing-Yun
QY
Sun
Yan-Li
YL
Ma
Na-Na
NN
Wang
Xiao-Yun
XY
Meng
Qing-Wei
QW
D002736:Chloroplasts / Q000201:enzymology*; D003545:Cysteine / Q000378:metabolism*; D004795:Enzyme Stability; D007700:Kinetics; D018551:Solanum lycopersicum / Q000201:enzymology*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D013696:Temperature
D050505:Mutant Proteins; D011994:Recombinant Proteins; D009247:NADH, NADPH Oxidoreductases; C023927:monodehydroascorbate reductase (NADH); D003545:Cysteine