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Molecules 2019, 24(4), 663; https://doi.org/10.3390/molecules24040663

A Convenient, Rapid, Sensitive, and Reliable Spectrophotometric Assay for Adenylate Kinase Activity

1
Biological Science Research Center, Southwest University, Beibei, Chongqing 400715, China
2
State Key Laboratory of Silkworm Genome Biology, College of Biotechnology, Southwest University, Beibei, Chongqing 400715, China
3
Chongqing Key Laboratory of Sericultural Science, Chongqing Engineering and Technology Research Center for Novel Silk Materials, Southwest University, Beibei, Chongqing 400715, China
*
Authors to whom correspondence should be addressed.
Academic Editor: Stefano Serra
Received: 19 January 2019 / Revised: 9 February 2019 / Accepted: 12 February 2019 / Published: 13 February 2019
(This article belongs to the Special Issue Enzymes, Biocatalysis and Chemical Biology)
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Abstract

Enzymatic activity assays are essential and critical for the study of enzyme kinetics. Adenylate kinase (Adk) plays a fundamental role in cellular energy and nucleotide homeostasis. To date, assays based on different principles have been used for the determination of Adk activity. Here, we show a spectrophotometric analysis technique to determine Adk activity with bromothymol blue as a pH indicator. We analyzed the effects of substrates and the pH indicator on the assay using orthogonal design and then established the most optimal assay for Adk activity. Subsequently, we evaluated the thermostability of Adk and the inhibitory effect of KCl on Adk activity with this assay. Our results show that this assay is simple, rapid, and precise. It shows great potential as an alternative to the conventional Adk activity assay. Our results also suggest that orthogonal design is an effective approach, which is very suitable for the optimization of complex enzyme reaction conditions. View Full-Text
Keywords: enzymatic activity assay; adenylate kinase; spectrophotometry; orthogonal experiment; bromothymol blue enzymatic activity assay; adenylate kinase; spectrophotometry; orthogonal experiment; bromothymol blue
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Song, K.; Wang, Y.; Li, Y.; Ding, C.; Cai, R.; Tao, G.; Zhao, P.; Xia, Q.; He, H. A Convenient, Rapid, Sensitive, and Reliable Spectrophotometric Assay for Adenylate Kinase Activity. Molecules 2019, 24, 663.

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