Pyridoxal 5'-phosphate dependent reactions: Analyzing the mechanism of aspartate aminotransferase.

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Mueser TC, Drago V, Kovalevsky A, Dajnowicz S

Pyridoxal 5'-phosphate dependent reactions: Analyzing the mechanism of aspartate aminotransferase.

Methods Enzymol. 2020;634:333-359. doi: 10.1016/bs.mie.2020.01.009. Epub 2020 Feb 13.

PubMed ID

32093839 [ View in PubMed

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Abstract

Enzyme catalysis is the primary activity in energy and information metabolism and enzyme cofactors are key to the catalytic ability of most enzymes. Pyridoxal 5'-phosphate (PLP) cofactor, derived from Vitamin B6, is widely distributed in nature and has significant latitude in catalytic diversity. X-ray crystallography has revealed the structures of diverse PLP dependent enzymes from multiple families. But these structures are incomplete, lacking the positions of protons essential for understanding enzymatic mechanisms. Here, we review the diversity of PLP and discuss the use of neutron crystallography and joint X-ray/neutron refinement of Fold Type I aspartate aminotransferase to visualize the positions of protons in both the internal and external aldimine forms. Strategies used to prepare extremely large crystals required for neutron diffraction and the approach to data refinement including the PLP cofactor are discussed. The observed positions of protons, including one located in a previously unknown low-barrier hydrogen bond, have been used to create more accurate models for computational analysis. The results revealed a new mechanism for the transaminase reaction where hyperconjugation is key to reducing the energy barrier which finally provides a clear explanation of the Dunathan alignment.

DrugBank Data that Cites this Article

Drug Targets

Drug	Target	Kind	Organism	Pharmacological Action	Actions
Pyridoxal phosphate	Aspartate aminotransferase (Protein Group)	Protein group	Humans	Unknown	Cofactor	Details