5ax8

Publication Abstract from PubMed

Mitochondrial aspartate aminotransferase (mAspAT) was recognized as a moonlighting enzyme because it has not only aminotransferase activity but also a high-affinity long-chain fatty acids (LCFA) binding site. This enzyme plays a key role in amino acid metabolism, biosynthesis of kynurenic acid and transport of the LCFA. Therefore, it is important to study the structure-function relationships of human mAspAT protein. In this work, the mature form of human mAspAT was expressed to a high level in Escherichia coli periplasmic space using pET-22b vector, purified by a combination of immobilized metal-affinity chromatography and cation exchange chromatography. Optimal activity of the enzyme occurred at a temperature of 47.5 masculineC and a pH of 8.5. Crystals of human mAspAT were grown using the hanging-drop vapour diffusion method at 277K with 0.1 M HEPES pH 6.8 and 25%(v/v) Jeffamine((R)) ED-2001 pH 6.8. The crystals diffracted to 2.99 A and belonged to the space group P1 with the unit-cell parameters a =56.7, b = 76.1, c = 94.2 A, alpha =78.0, beta =85.6, gamma = 78.4 masculine. Elucidation of mAspAT structure can provide a molecular basis towards understanding catalysis mechanism and substrate binding site of enzyme.

Recombinant expression, purification and crystallographic studies of the mature form of human mitochondrial aspartate aminotransferase.,Jiang X, Wang J, Chang H, Zhou Y Biosci Trends. 2016 Mar 10;10(1):79-84. doi: 10.5582/bst.2015.01150. Epub 2016, Feb 22. PMID:26902786^[2]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.