Mutagenesis of the l-Amino Acid Ligase RizA Increased the Production of Bioactive Dipeptides

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Bordewick, S.; Berger, R.G.; Ersoy, F.: Mutagenesis of the l-Amino Acid Ligase RizA Increased the Production of Bioactive Dipeptides. In: Catalysts : open access journal 11 (2021), Nr. 11, 1385. DOI:

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The l-amino acid ligase RizA from B. subtilis selectively synthesizes dipeptides containing an N-terminal arginine. Many arginyl dipeptides have salt-taste enhancing properties while Arg-Phe has been found to have an antihypertensive effect. A total of 21 RizA variants were created by site-directed mutagenesis of eight amino acids in the substrate binding pocket. The variants were recombinantly produced in E. coli and purified by affinity chromatography. Biocatalytic reactions were set up with arginine and four amino acids differing in size and polarity (aspartic acid, serine, alanine, and phenylalanine) and were analyzed by RP-HPLC with fluorescence detection. Variant T81F significantly improved the yield in comparison to wild type RizA for aspartic acid (7 to 17%), serine (33 to 47%) and alanine (12 to 17%). S84F increased product yield similarly for aspartic acid (7 to 17%) and serine (33 to 42%). D376E increased the yield with alanine (12 to 19%) and phenylalanine (11 to 26%). The largest change was observed for S156A, which showed a yield for Arg-Phe of 40% corresponding to a 270% increase in product concentration. This study expands the knowledge about positions governing the substrate specificity of RizA and may help to inform future protein engineering endeavors.
License of this version: CC BY 4.0 Unported
Document Type: Article
Publishing status: publishedVersion
Issue Date: 2021
Appears in Collections:Naturwissenschaftliche Fakultät

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1 image of flag of Germany Germany 59 30.57%
2 image of flag of United States United States 50 25.91%
3 image of flag of United Kingdom United Kingdom 17 8.81%
4 image of flag of No geo information available No geo information available 10 5.18%
5 image of flag of Singapore Singapore 10 5.18%
6 image of flag of Russian Federation Russian Federation 6 3.11%
7 image of flag of Japan Japan 6 3.11%
8 image of flag of United Arab Emirates United Arab Emirates 5 2.59%
9 image of flag of China China 4 2.07%
10 image of flag of Belgium Belgium 3 1.55%
    other countries 23 11.92%

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