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Determinants of the Nucleotide Specificity in the Carbohydrate Epimerase Family 1
Author(s) -
Van Overtveldt Stevie,
Da Costa Matthieu,
Gevaert Ophelia,
Joosten HenkJan,
Beerens Koen,
Desmet Tom
Publication year - 2020
Publication title -
biotechnology journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.144
H-Index - 84
eISSN - 1860-7314
pISSN - 1860-6768
DOI - 10.1002/biot.202000132
Subject(s) - thermus thermophilus , nucleotide , biochemistry , carbohydrate , nucleotide sugar , enzyme , thermus , biology , chemistry , gene , escherichia coli , thermophile
In recent years, carbohydrate epimerases have attracted increasing attention as promising biocatalysts for the production of specialty sugars and derivatives. The vast majority of these enzymes are active on nucleotide‐activated sugars, rather than on their free counterparts. Although such epimerases are known to have a clear preference for a particular nucleotide (UDP, GDP, CDP, or ADP), very little is known about the determinants of the respective specificities. In this work, sequence motifs are identified that correlate with the different nucleotide specificities in one of the main epimerase superfamilies, carbohydrate epimerase 1 (CEP1). To confirm their relevance, GDP‐ and CDP‐specific residues are introduced into the UDP‐glucose 4‐epimerase from Thermus thermophilus , resulting in a 3‐fold and 13‐fold reduction in K M for GDP‐Glc and CDP‐Glc, respectively. Moreover, several variants are severely crippled in UDP‐Glc activity, which further underlines the crucial role of the identified positions. Hence, the analysis should prove to be valuable for the further exploration and application of epimerases involved in carbohydrate synthesis.