Mechanistic and Structural Insight to an Evolved Benzoylformate Decarboxylase with Enhanced Pyruvate Decarboxylase Activity
AbstractBenzoylformate decarboxylase (BFDC) and pyruvate decarboxylase (PDC) are thiamin diphosphate-dependent enzymes that share some structural and mechanistic similarities. Both enzymes catalyze the nonoxidative decarboxylation of 2-keto acids, yet differ considerably in their substrate specificity. In particular, the BFDC from P. putida exhibits very limited activity with pyruvate, whereas the PDCs from S. cerevisiae or from Z. mobilis show virtually no activity with benzoylformate (phenylglyoxylate). Previously, saturation mutagenesis was used to generate the BFDC T377L/A460Y variant, which exhibited a greater than 10,000-fold increase in pyruvate/benzoylformate substrate utilization ratio compared to that of wtBFDC. Much of this change could be attributed to an improvement in the Km value for pyruvate and, concomitantly, a decrease in the kcat value for benzoylformate. However, the steady-state data did not provide any details about changes in individual catalytic steps. To gain insight into the changes in conversion rates of pyruvate and benzoylformate to acetaldehyde and benzaldehyde, respectively, by the BFDC T377L/A460Y variant, reaction intermediates of both substrates were analyzed by NMR and microscopic rate constants for the elementary catalytic steps were calculated. Herein we also report the high resolution X-ray structure of the BFDC T377L/A460Y variant, which provides context for the observed changes in substrate specificity. View Full-Text
Scifeed alert for new publicationsNever miss any articles matching your research from any publisher
- Get alerts for new papers matching your research
- Find out the new papers from selected authors
- Updated daily for 49'000+ journals and 6000+ publishers
- Define your Scifeed now
Andrews, F.H.; Wechsler, C.; Rogers, M.P.; Meyer, D.; Tittmann, K.; McLeish, M.J. Mechanistic and Structural Insight to an Evolved Benzoylformate Decarboxylase with Enhanced Pyruvate Decarboxylase Activity. Catalysts 2016, 6, 190.
Andrews FH, Wechsler C, Rogers MP, Meyer D, Tittmann K, McLeish MJ. Mechanistic and Structural Insight to an Evolved Benzoylformate Decarboxylase with Enhanced Pyruvate Decarboxylase Activity. Catalysts. 2016; 6(12):190.Chicago/Turabian Style
Andrews, Forest H.; Wechsler, Cindy; Rogers, Megan P.; Meyer, Danilo; Tittmann, Kai; McLeish, Michael J. 2016. "Mechanistic and Structural Insight to an Evolved Benzoylformate Decarboxylase with Enhanced Pyruvate Decarboxylase Activity." Catalysts 6, no. 12: 190.
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.