Conserved calcium-binding residues at the ca-i site involved in fructooligosaccharide synthesis by lactobacillus reuteri 121 inulosucrase

Charoenwongpaiboon, Thanapon, Punnatin, Panachai, Klaewkla, Methus, Pramoj Na Ayutthaya, Pratchaya, Wangpaiboon, Karan, Chunsrivirot, Surasak, Field, Robert A. ORCID: https://orcid.org/0000-0001-8574-0275 and Pichyangkura, Rath (2020) Conserved calcium-binding residues at the ca-i site involved in fructooligosaccharide synthesis by lactobacillus reuteri 121 inulosucrase. ACS Omega, 5 (43). pp. 28001-28011. ISSN 2470-1343

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Abstract

Inulosucrase is an enzyme that synthesizes inulin-type β-2,1-linked fructooligosaccharides (IFOS) from sucrose. Previous studies have shown that calcium is important for the activity and stability of Lactobacillus reuteri 121 inulosucrase (LrInu). Here, mutational analyses of four conserved calcium-binding site I (Ca-I) residues of LrInu, Asp418, Gln449, Asn488, and Asp520 were performed. Alanine substitution for these residues not only reduced the stability and activity of LrInu, but also modulated the pattern of the IFOS produced. Circular dichroism spectroscopy and molecular dynamics simulation indicated that these mutations had limited impact on the overall conformation of the enzyme. One of Ca-I residues most critical for controlling LrInu-mediated polymerization of IFOS, Asp418, was also subjected to mutagenesis, generating D418E, D418H, D418L, D418N, D418S, and D418W. The activity of these mutants demonstrated that the IFOS chain length could be controlled by a single mutation at the Ca-I site.

Item Type:	Article
Additional Information:	Funding Information: T.C. is thankful to Department of Biochemistry, Faculty of Science, Chulalongkorn University and Department of Biological Chemistry, John Innes Centre. S.C. was partially supported by the Structural and Computational Biology Research Unit, Faculty of Science, Rachadaphiseksomphot Endowment Fund, Chulalongkorn University. Publisher Copyright: ©
Uncontrolled Keywords:	chemistry(all),chemical engineering(all) ,/dk/atira/pure/subjectarea/asjc/1600
Faculty \ School:	Faculty of Science > School of Chemistry, Pharmacy and Pharmacology
Related URLs:	http://www.scopus.com/inward/record.url?...
Depositing User:	LivePure Connector
Date Deposited:	02 Sep 2024 13:31
Last Modified:	25 Sep 2024 18:05
URI:	https://ueaeprints.uea.ac.uk/id/eprint/96427
DOI:	10.1021/acsomega.0c03521

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