Structural Modelling and Structure-Function Analysis of <i>Zymomonas mobilis</i> Levansucrase

Abstract

Levansucrases are bacterial enzymes which produce fructan polymers from sucrose via hydrolysis and transfructosylation activities. These polymers; levan and fructooligosaccharides are valuable for food and pharmaceutical industries. Levansucrases from Gram-positive bacteria such as Bacillus subtilis tend to produce levan, while those from Gram-negative bacteria preferentially produce fructooligosaccharides. Zymomonas mobilis is an efficient levansucrase producer and its extracellular levansucrase can produce both fructooligosaccharides and levan depending on the reaction parameters. In this study, the structure of Z. mobilis levansucrase was modeled in order to help to understand the structure-function relationship of the enzyme. Furthermore, amino acids previously reported to be important for levansucrase activity were mapped on the model. The structural model presents a five-bladed propeller with a deep, negatively charged central pocket, similar to other bacterial levansucrases. Mapping showed that amino acids which previously reported to affect fructan length are located on the periphery of the structure covering the active site central pocket. Thus it is showed that, for the first time, that hydrolysis and transfructosylation reactions are catalyzed on different parts of Z. mobilis levansucrase structure. The structural location of the critical amino acids will pave the way to identify other residues which control fructan length by site directed mutagenesis without altering the overall fold of the enzyme.

Lscβ and lscγ, two novel levansucrases of Pseudomonas syringae pv. actinidiae biovar 3, the causal agent of bacterial canker of kiwifruit, show different enzymatic properties

“…To the best of our knowledge, this is the first report with experimental evidence highlighting the possible different localization of the hydrolysis and transfructosylation reactions in lsc which till now were only supposed by molecular modelling [57].…”

A novel fold for acyltransferase-3 (AT3) proteins provides a framework for transmembrane acyl-group transfer

“…The structure of OafB was predicted by RaptorX ( Wang et al, 2016 ; Källberg et al, 2014 ; Ma et al, 2015 ) using the protein sequence of a Salmonella rhamnose O-acetyltransferase (OafB). This method has been successfully validated with numerous proteins ( Sharma et al, 2021 ; Mariani et al, 2011 ; Lopes-Rodrigues et al, 2019 ; Bakar and Kaplan-türköz, 2017 ; Xu and Wang, 2019 ). The structure of OafB consists of two key domains: the AT3 domain and SGNH domain ( Figure 1C ).…”

Tailoring fructooligosaccharides composition with engineered Zymomonas mobilis ZM4

“…Cys121 and Cys151 affect the structure of the protein and consequently the enzyme activity. On the other hand, Cys242 is not related with protein folding but affects the final product type (Li et al 2008;Kaplan-Turkoz and Bakar 2017). Although the heterologous production of FOS by different organisms has proven to be a successful strategy to optimize and change the FOS composition (Zhang et al 2017;Hernández et al 2018;Ko et al 2019;Rodrigo-Frutos et al 2019), there are no reports regarding the heterologous production of FOS by Z. mobilis.…”