Evaluation of different pretreatment protocols to detect accurately clinical carbapenemase-producing Enterobacteriaceae by MALDI-TOF.

Objectives: Carbapenemase-resistant bacteria are increasingly spreading worldwide causing public concern due to their ability to elude antimicrobial treatment. Early identification of these bacteria is therefore of high importance. Here, we describe the development of a simple and robust protocol for the detection of carbapenemase activity in clinical isolates of Enterobacteriaceae, suitable for routine and clinical applications.

Homogeneity and heterogeneity in amylase production by Bacillus subtilis under different growth conditions.

Background: Bacillus subtilis is an important cell factory for the biotechnological industry due to its ability to secrete commercially relevant proteins in large amounts directly into the growth medium. However, hyper-secretion of proteins, such as α-amylases, leads to induction of the secretion stress-responsive CssR-CssS regulatory system, resulting in up-regulation of the HtrA and HtrB proteases. These proteases degrade misfolded proteins secreted via the Sec pathway, resulting in a loss of product.

A mutation leading to super-assembly of twin-arginine translocase (Tat) protein complexes.

The Tat system transports folded proteins across the bacterial plasma membrane. The mechanism is believed to involve coalescence of a TatC-containing unit with a separate TatA complex, but the full translocation complex has never been visualised and the assembly process is poorly defined. We report the analysis of the Bacillus subtilis TatAyCy system, which occurs as separate TatAyCy and TatAy complexes at steady state, using single-particle electron microscopy (EM) and advanced atomic force microscopy (AFM) approaches.

Co-factor insertion and disulfide bond requirements for twin-arginine translocase-dependent export of the Bacillus subtilis Rieske protein QcrA.

The twin-arginine translocation (Tat) pathway can transport folded and co-factor-containing cargo proteins over bacterial cytoplasmic membranes. Functional Tat machinery components, a folded state of the cargo protein and correct co-factor insertion in the cargo protein are generally considered as prerequisites for successful translocation. The present studies were aimed at a dissection of these requirements with regard to the Rieske iron-sulfur protein QcrA of Bacillus subtilis.

Degradation of extracytoplasmic catalysts for protein folding in Bacillus subtilis.

The general protein secretion pathway of Bacillus subtilis has a high capacity for protein export from the cytoplasm, which is exploited in the biotechnological production of a wide range of enzymes. These exported proteins pass the membrane in an unfolded state, and accordingly, they have to fold into their active and protease-resistant conformations once membrane passage is completed. The lipoprotein PrsA and the membrane proteins HtrA and HtrB facilitate the extracytoplasmic folding and quality control of exported proteins.

The Tat system of Gram-positive bacteria.

The twin-arginine protein translocation (Tat) system has a unique ability to translocate folded and co-factor-containing proteins across lipid bilayers. The Tat pathway is present in bacteria, archaea and in the thylakoid membranes of chloroplasts and, depending on the organism and environmental conditions, it can be deemed important for cell survival, virulence or bioproduction. This review provides an overview of the current understanding of the Tat system with specific focus on Gram-positive bacteria.

The Bacillus subtilis EfeUOB transporter is essential for high-affinity acquisition of ferrous and ferric iron.

Efficient uptake of iron is of critical importance for growth and viability of microbial cells. Nevertheless, several mechanisms for iron uptake are not yet clearly defined. Here we report that the widely conserved transporter EfeUOB employs an unprecedented dual-mode mechanism for acquisition of ferrous (Fe[II]) and ferric (Fe[III]) iron in the bacterium Bacillus subtilis.

Ultrastructural characterisation of Bacillus subtilis TatA complexes suggests they are too small to form homooligomeric translocation pores.

Tat-dependent protein transport permits the traffic of fully folded proteins across membranes in bacteria and chloroplasts. The mechanism by which this occurs is not understood. Current theories propose that a key step requires the coalescence of a substrate-binding TatC-containing complex with a TatA complex, which forms pores of varying sizes that could accommodate different substrates.

Mapping the twin-arginine protein translocation network of Bacillus subtilis.

Bacteria employ twin-arginine translocation (Tat) pathways for the transport of folded proteins to extracytoplasmic destinations. In recent years, most studies on bacterial Tat pathways addressed the membrane-bound TatA(B)C subunits of the Tat translocase, and the specific interactions between this translocase and its substrate proteins. In contrast, relatively few studies investigated possible coactors in the TatA(B)C-dependent protein translocation process.

High-salinity growth conditions promote Tat-independent secretion of Tat substrates in Bacillus subtilis.

The Gram-positive bacterium Bacillus subtilis contains two Tat translocases, which can facilitate transport of folded proteins across the plasma membrane. Previous research has shown that Tat-dependent protein secretion in B. subtilis is a highly selective process and that heterologous proteins, such as the green fluorescent protein (GFP), are poor Tat substrates in this organism.

Degradation of the twin-arginine translocation substrate YwbN by extracytoplasmic proteases of Bacillus subtilis.

Bacterial twin-arginine translocases can export fully folded proteins from the cytoplasm. Such proteins are usually resistant to proteolysis. Here we show that multiple extracellular proteases degrade the B.

Specific targeting of the metallophosphoesterase YkuE to the bacillus cell wall requires the twin-arginine translocation system.

The twin-arginine translocation (Tat) pathway is dedicated to the transport of fully folded proteins across the cytoplasmic membranes of many bacteria and the chloroplast thylakoidal membrane. Accordingly, Tat-dependently translocated proteins are known to be delivered to the periplasm of Gram-negative bacteria, the growth medium of Gram-positive bacteria, and the thylakoid lumen. Here, we present the first example of a protein, YkuE of Bacillus subtilis, that is specifically targeted by the Tat pathway to the cell wall of a Gram-positive bacterium.

TatAc, the third TatA subunit of Bacillus subtilis, can form active twin-arginine translocases with the TatCd and TatCy subunits.

Two independent twin-arginine translocases (Tat) for protein secretion were previously identified in the Gram-positive bacterium Bacillus subtilis. These consist of the TatAd-TatCd and TatAy-TatCy subunits. The function of a third TatA subunit named TatAc was unknown.

Environmental salinity determines the specificity and need for Tat-dependent secretion of the YwbN protein in Bacillus subtilis.

Twin-arginine protein translocation (Tat) pathways are required for transport of folded proteins across bacterial, archaeal and chloroplast membranes. Recent studies indicate that Tat has evolved into a mainstream pathway for protein secretion in certain halophilic archaea, which thrive in highly saline environments. Here, we investigated the effects of environmental salinity on Tat-dependent protein secretion by the Gram-positive soil bacterium Bacillus subtilis, which encounters widely differing salt concentrations in its natural habitats.