Influence of Tags on the Binding Affinity of Acyl-CoA Binding Protein
Jizhong Han, Yunlong Sun, Yu Chen, Haoran Li, Mengmeng Liu and Bin Zeng
American Journal of Biochemistry and Biotechnology
Tagged fusion proteins are frequently employed for protein purification methods, but their effects on protein function and binding affinity are rarely studied. Here we expressed recombinant protein Acyl-CoA Binding Protein (ACBP) cloned from the full-length cDNA of Aspergillus oryzae and Saccharomyces cerevisiae. ACBP was expressed in Escherichia coli fused to a Maltose-Binding Protein (MBP) and Histidine-tag fusion. Recombinant ACBP was purified using affinity chromatography columns and high protein purity was achieved. Microscale Thermophoresis (MST) binding assays showed that recombinant AoAcbp1 had a greater affinity for Palmitoyl-CoA (Kd = 35 nM) and Stearoyl-CoA (Kd = 23 nM) whilst recombinant ScAcbp had a greater affinity for Myristoyl-CoA (Kd = 31 nM) and Palmitoyl-CoA (Kd = 51 nM). In addition, MBP tagged ACBP had comparable binding affinities to His-tagged ACBP. Taken together, these data highlight that the size of the tagged fusion protein does not influence protein ACBP binding affinity.
© 2019 Jizhong Han, Yunlong Sun, Yu Chen, Haoran Li, Mengmeng Liu and Bin Zeng. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.