Analysis of hydrolytic differences of free and "polyacrylic acid (PAAc)-conjugated trypsin and chymotrypsin" by using fluorescence lifetime distributions.

Preparative Biochemistry & Biotechnology
Ümmügülsüm PolatEmine Karakuş

Abstract

Electroanalytic, photometric or fluorometric methods may provide information about the presence of proteolysis in a related sample, but they cannot accurately identify which protease belongs to the proteolytic activity. In other words, they cannot distinguish the proteases according to the differences in their activities. Previous studies on the chymotrypsin and trypsin have shown that the activities of proteases can be distinguished from each other by fluorescence lifetime distributions. In this study, it is aimed to show the sensitivity of the distributional model on the proteolytic activities of the two proteases. For this purpose, the proteolytic activities have been reduced by making covalent conjugation with polyacrylic acid (PAAc) and the effects of the low activities were examined on Bovine Serum Albumin (BSA) which excimer emission character was incorporated into its structure by modification with N-(1-pyrenyl)maleimide. The time-resolved spectrofluorometer was used to collect fluorescence decay data at λ(excimer) = 464 nm, which were analyzed by using Exponential Series Method (ESM) to obtain the changes of lifetime distributions. The results showed a significant decline in the activities. Despite the very low activit...Continue Reading

References

Jun 29, 1976·Biochemistry·C W Wu, L R Yarbrough
May 6, 1998·Protein Science : a Publication of the Protein Society·A R Khan, M N James
Nov 30, 2004·Journal of Colloid and Interface Science·Shengcong LiufuYuping Li
Jul 5, 2012·Methods in Molecular Biology·Sarah SchreursYves Engelborghs
Jul 11, 2013·Bioconjugate Chemistry·Mohammad RashidianMark D Distefano
Oct 23, 2015·Spectrochimica Acta. Part A, Molecular and Biomolecular Spectroscopy·İbrahim Ethem ÖzyiğitÖnder Pekcan

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Citations

Sep 17, 2020·Preparative Biochemistry & Biotechnology·Yavuz Selim ToksözEmine Karakuş

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Methods Mentioned

BETA
gel filtration
Fluorescence

Software Mentioned

PTI FeliX32
ESM

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