A study of targeted mutation of l-rhamnose isomerase to improve the conversion efficiency of D-allose

Duan, Shuangshuang; Chen, Yonghua; Wang, Guodong; Li, Zebin; Dong, Shouliang; Wu, Yufan; Wang, Yuanwei; Ma, Chunling; Wang, Ruiming

doi:10.1016/j.enzmictec.2023.110259

Cited by 3 publications

(1 citation statement)

References 27 publications

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“…However, the enzyme activity of L-RhIs on D-allose is relatively low for industrial usage ( Pseudomonas stutzeri L-RhI, 7.5 U/mg, Leang et al 2004b ; Mesorhizobium loti L-RhI, 3.0 U/mg, Takata et al 2011 ; Bacillus pallidus L-RhI, 2.6 U/mg, Poonperm et al 2007 ; Caldicellulosiruptor obsidiansis , L-RhI, 13.7 U/mg, Chen et al 2018a ; Thermobacillus composti L-RhI, 1.7 U/mg; Thermotoga maritima L-RhI, 6.7 U/mg, Park et al 2010 ; Thermoanaerobacterium saccharolyticum L-RhI, 5.7 U/mg, Lin et al 2010 ). Thus, several studies seeking to alter the substrate specificity or improve activity toward D-allose have been conducted (Tseng et al 2022 ; Chen et al 2018b ; Duan et al 2023 ).…”

Section: Introductionmentioning

confidence: 99%

X-ray structure and characterization of a probiotic Lactobacillus rhamnosus Probio-M9 L-rhamnose isomerase

Yoshida,

Yamamoto,

Kurahara

et al. 2024

Appl Microbiol Biotechnol

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A recombinant L-rhamnose isomerase (L-RhI) from probiotic Lactobacillus rhamnosus Probio-M9 (L. rhamnosus Probio-M9) was expressed. L. rhamnosus Probio-M9 was isolated from human colostrum and identified as a probiotic lactic acid bacterium, which can grow using L-rhamnose. L-RhI is one of the enzymes involved in L-rhamnose metabolism and catalyzes the reversible isomerization between L-rhamnose and L-rhamnulose. Some L-RhIs were reported to catalyze isomerization not only between L-rhamnose and L-rhamnulose but also between D-allulose and D-allose, which are known as rare sugars. Those L-RhIs are attractive enzymes for rare sugar production and have the potential to be further improved by enzyme engineering; however, the known crystal structures of L-RhIs recognizing rare sugars are limited. In addition, the optimum pH levels of most reported L-RhIs are basic rather than neutral, and such a basic condition causes non-enzymatic aldose-ketose isomerization, resulting in unexpected by-products. Herein, we report the crystal structures of L. rhamnosus Probio-M9 L-RhI (LrL-RhI) in complexes with L-rhamnose, D-allulose, and D-allose, which show enzyme activity toward L-rhamnose, D-allulose, and D-allose in acidic conditions, though the activity toward D-allose was low. In the complex with L-rhamnose, L-rhamnopyranose was found in the catalytic site, showing favorable recognition for catalysis. In the complex with D-allulose, D-allulofuranose and ring-opened D-allulose were observed in the catalytic site. However, bound D-allose in the pyranose form was found in the catalytic site of the complex with D-allose, which was unfavorable for recognition, like an inhibition mode. The structure of the complex may explain the low activity toward D-allose. Key points • Crystal structures of LrL-RhI in complexes with substrates were determined. • LrL-RhI exhibits enzyme activity toward L-rhamnose, D-allulose, and D-allose. • The LrL-RhI is active in acidic conditions.

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