Functional Characterization of Transporters for L-Aspartate in Bacillus licheniformis

Hanrong Wang,Zhenghua Gu,Zhongyang Ding,Yupeng Zhang,Fengxu Xiao,Youran Li,Sha Xu,Guiyang Shi,Liang Zhang

doi:10.3390/fermentation8010022

Abstract

Amino acid efflux and influx transport systems play vital roles in industrial microorganisms’ cell growth and metabolism. However, although biochemically characterized, most of them remain unknown at the molecular level in Bacillus licheniformis. In this study, three proteins, namely, YdgF, YvbW, and YveA, were predicted to be involved in the active transport of L-aspartate (L-Asp). This was verified by manipulating their encoding genes. When growing in the minimal medium with L-Asp as the only carbon and nitrogen source, the growth of strains lacking proteins YdgF, YvbW, and YveA was significantly inhibited compared with the wild-type strains, while supplementing the expression of the corresponding proteins in the single-gene knockout strains could alleviate the inhibition. Upon overexpression, the recombinant proteins mediated the accumulation of L-aspartate to varying degrees. Compared with the wild-type strains, the single knockout strains of the three protein genes exhibited reduced absorption of L-aspartate. In addition, this study focused on the effects of these three proteins on the absorption of β-alanine, L-glutamate, D-serine, D-alanine, and glycine.

Highlights

Published: 7 January 2022Amino acids are widely used in food, feed, medicine, and cosmetics, and are mainly produced by chemical synthesis, enzyme catalysis, or a combination of the two
This study focused on the transport of L-Asp and other related amino acids by three types of amino acid permeases on the cell membrane of B. licheniformis
Since the functions of the amino acid permeases YdgF, YvbW, and YveA are still not clear, these three proteins were selected as the research objects, where their encoding genes are designated as ydgF, yvbW, and yveA, respectively

Summary

Introduction

Amino acids are widely used in food, feed, medicine, and cosmetics, and are mainly produced by chemical synthesis, enzyme catalysis, or a combination of the two. High concentrations of amino acids inside or outside the cell are often not conducive to fermentation production [3,4]. Some amino-acid-producing enzymes are inhibited by the substrate; for instance, L-aspartic acid α-decarboxylase can be irreversibly inhibited by L-aspartic acid. Excessive intracellular accumulation of certain amino acids will reduce fermentation production and have a toxic effect on cells. High intracellular cysteine concentration can lead to DNA damage of the host and have a toxic effect on the producing strain. In the production of alanine fermentation, the high content of intracellular alanine will inhibit cell growth [6].

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Results

Conclusion