Hemoglobin from the blood clam Tegillarca granosa (Tg-HbIIA, Tg-HbIIB): expression and antibacterial activity of recombinant proteins

Abstract

The hemoglobins produced by Tegillarca granosa have antibacterial activity toward some Gram-positive and Gram-negative bacteria. In this study, the genes encoding the recombinant proteins Tg-HbIIA and Tg-HbIIB were cloned from T. granosa hemocytes by RT-PCR, and the proteins were expressed in Escherichia coli Transetta (DE3). The proteins were purified using a HisTrap FF affinity chromatography column under denaturing conditions and refolded at 4 °C by urea gradient dialysis, and the antibacterial activity of the recombinant proteins was determined. The Tg-HbIIA protein had antibacterial activity toward Vibrio harveyi and Pseudomonas putida, with the minimum inhibition concentration (MIC) values of 65.8 ug/ml and 4.11 ug/ml, respectively. The Tg-HbIIB protein had antibacterial activity toward V. harveyi, P. putida and Acinetobacter baumanii, with MIC values of 158 ug/ml, 39.5 ug/ml and 79 ug/ml, respectively. They had no antibacterial activity against Staphylococcus aureus, E. coli, B. firmus, B. subtilis, S. epidermidis, or V. parahaemolyticus. This study provides a basis for further research on the antibacterial function and mechanism of hemoglobin.