Intrinsic Properties Analysis of Multiproteases System from Marine Bacteria by Inhibitor-Subsatrate Immersion Zymography

摘要

Based on digital image analysis techniques and inhibitor-substrate immersing zymography, intrinsic properties of each active component in the enzymatic system secreted by marine bacteria were studied. This method provides an easy way to characterize the proteases in situ, which can be further verified by Mass spectrometry. Compared to the Folin phenol method, a traditional method used to determine proteases activities, the inhibitor-substrate immersing zymography method coupled with digital image analysis used in this study could determine caseinolytic activity and measure gelatinolytic activity at the same time. The effect on activities of extracellular proteases by inhibitor (phenylmethylsulfonyl fluoride or 1, 10-Phenanthroline) can be quantified by gray value changes of the corresponding band after electrophoretic separation. Because of its high throughput, great sensitivity, and convenient operation, inhibitor-substrate immersing zymography can be used to demonstrate the natural diversity of protein hydrolases and multienzyme expression systems. Thus, it is an effective approach to study the functional proteomics of proteases secreted by marine bacteria.