Limited proteolysis improves E.coli Hfq crystal structure resolution

Hfq is an abundant RNA chaperone in bacteria. Hfq facilitates sRNAs targeting their antisense RNAs that regulates bacteria responses in stressing environments. Hfq is a homohexameric protein which has six histidine residues on the surface of its inner ring. Therefore,nickel affinity chromatography can be used to directly purify this protein from E. coli. After size exclusion chromatography,the purity of this protein achieved 70% . Before crystallization,this protein was limited proteolysised by chymotrypsin to cut C-terminal flexible loop. The digested Hfq fragment was crystallized in many conditions containing ( NH4 ) 2SO4 as precipitant. But the initial needle-like crystals are too small to be used for crystal data collection. After crystallization optimization,bigger crystals grew in two conditions. Finally,two Hfq crystal structures were solved. The resolution of one crystal structure was 1. 63 .