Recombinant expression, purification, and characterization of an acyl-CoA binding protein from Aspergillus oryzae
2016
Hao, Qing | Liu, Xiaoguang | Zhao, Guozhong | Jiang, Lu | Li, Ming | Zeng, Bin
OBJECTIVES: To characterize biochemically the lipid metabolism-regulating acyl-CoA binding protein (ACBP) from the industrially-important fungus Aspergillus oryzae. RESULTS: A full-length cDNA encoding a candidate ACBP from A. oryzae (AoACBP) was cloned and expressed in Escherichia coli as a maltose-binding protein (MBP) fusion protein. The MBP-AoACBP protein was purified by an amylose resin chromatography column. SDS-PAGE showed that MBP-AoACBP has an estimated molecular weight of 82 kDa. Microscale thermophoresis binding assay showed that the recombinant AoACBP displayed much greater affinity for palmitoyl-CoA (K d = 80 nM) than for myristoyl-CoA (K d = 510 nM), thus demonstrating the preference of AoACBP for long-chain acyl-CoA. CONCLUSION: The data support the identification of AoACBP as a long-chain ACBP in A. oryzae.
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