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.
Keywords: Acyl-CoA binding affinity; Acyl-CoA binding protein; Aspergillus oryzae; Microscale thermophoresis binding assay; Myristoyl-CoA; Palmitoyl-CoA.