Three Oligopeptide-binding Proteins Are Involved in the Oligopeptide Transport of Streptococcus thermophilus

Peggy Garault,Dominique Le Bars,Colette Besset,Véronique Monnet

doi:10.1074/jbc.m107002200

Abstract

The functions necessary for bacterial growth strongly depend on the features of the bacteria and the components of the growth media. Our objective was to identify the functions essential to the optimum growth of Streptococcus thermophilus in milk. Using random insertional mutagenesis on a S. thermophilus strain chosen for its ability to grow rapidly in milk, we obtained several mutants incapable of rapid growth in milk. We isolated and characterized one of these mutants in which an amiA1 gene encoding an oligopeptide-binding protein (OBP) was interrupted. This gene was a part of an operon containing all the components of an ATP binding cassette transporter. Three highly homologous amiA genes encoding OBPs work with the same components of the ATP transport system. Their simultaneous inactivation led to a drastic diminution in the growth rate in milk and the absence of growth in chemically defined medium containing peptides as the nitrogen source. We constructed single and multiple negative mutants for AmiAs and cell wall proteinase (PrtS), the only proteinase capable of hydrolyzing casein oligopeptides outside the cell. Growth experiments in chemically defined medium containing peptides indicated that AmiA1, AmiA2, and AmiA3 exhibited overlapping substrate specificities, and that the whole system allows the transport of peptides containing from 3 to 23 residues.

Highlights

Oligopeptide transport systems are key channels between the environment and the inner part of micro-organisms, which have been described in numerous Gram-negative and Grampositive bacteria. They generally internalize peptides with an ATP-driving force and belong to the ABC transporter family [1]. They are composed of oligopeptide-binding proteins (OBP),1 which are periplasmic in Gram-negative bacteria and membrane-associated in Gram-positive bacteria; transmembrane proteins that form a channel for the passage of oligopeptides and inner membrane-associated ATPases, which provide the energy for transport
In the case of lactic acid bacteria, which are auxotrophic for several amino acids (8 –10) and which are used to growing in milk, a medium containing a low level of free amino acids [11], the oligopeptide transport system is fundamental to optimal growth
Oligopeptide Uptake Is Essential to the Growth of S. thermophilus in Peptide-containing Media—In addition to cell wall protease and the purine and branched-chain amino acid biosynthesis pathways, the oligopeptide transport system is one of the functions necessary for the optimum growth of S. thermophilus in milk [26, 24, 35]

Summary

Introduction

Oligopeptide transport systems are key channels between the environment and the inner part of micro-organisms, which have been described in numerous Gram-negative and Grampositive bacteria. They generally internalize peptides with an ATP-driving force and belong to the ABC transporter family [1]. We have identified and characterized the oligopeptide transport system of S. thermophilus We demonstrate that it works with three functional oligopeptide-binding proteins, that it is capable of transporting entities as large as 23 amino acid peptides, and that it plays a major role in nutrition

Objectives

Results

Conclusion