Preface

AcrB is one of the many important multi-drug transporters found in a Gram-negative bacteria such as E. coli. The primary function of AcrB enables Gram-negative bacteria to exhibit intrinsic tolerance toward multiple antibiotic drugs. We are beginning to understand how this ability to recognise a variety of drugs is expressed by AcrB. Here, we will explore this ability using the basis of molecular weight differentiation adopted by AcrB for the regonition of antibiotic drugs and their subsequent transportation out of the cell.

In previous studies performed with AcrB, low molecular weight (MW) drugs such as minocycline & doxorubicin have been shown to bind to a specific phenylalanine cluster region (the distal pocket) of the binding monomer of AcrB. Further analysis revealed the drugs were exported out of E. coli via a 3-step functionally-rotating mechanism*.

The latest studies involving the broad specificity range of AcrB has revealed an additional binding pocket in the access monomer of the exporter molecule which is suspected to bind high molecular weight drugs such as rifampicin and erythromycin. It is this additional binding site that will be the key focus of our analysis of AcrB as we compare it to the previously studied distal pocket binding site.

*Further elaboration on the mechanism can be found under 'Structure' (Proposed Transport Mechanisms- An Overview) and 'Functional Mechanism'.

A brief preview of the AcrB multi-drug exporter:

A homotrimer component of a tripartite complex - with the AcrA membrane fusion protein which connects AcrB to TolC, the outer membrane channel which allows drugs to bypass the outer membrane barrier of bacteria and straight into the external medium.
Tripartite system is also know as the Efflux pump complex which may be energy dependent to perform its functions.
Previous experiment with deletion of genes encoding for one or many components of the AcrB-AcrA-TolC tripartite system showed increased antibiotic susceptibility in E. coli.

This video here illustrates the basic principles behind the efflux pump function mechanism such as the one used by AcrB.

Basic function of an efflux pump

Overview

Gram-negative bacteria exhibit higher intrinsic levels of resistance to a range of drugs as compared to the Gram-positive strain. This enhanced tolerance is attributed to multidrug efflux pumps embedded in cell membranes which recognize a wide variety of substrates. These transporters are tripartite and associate with periplasmic proteins to form complexes responsible for the efflux of drugs out of the cell directly to the external surroundings.

Our blog is concentrated on a particular transporter called AcrB, which associates with AcrA and TolC. The primary focus is the mechanism utilized by AcrB to transport both high and low molecular weight drugs. Crystal structures of the protein have revealed and provided evidence that the presence of an additional binding site facilitates the mechanism. Results from site-directed mutagenesis have contributed affirmation toward the proposed mechanism.

Structural, mechanistic and experimental details relevant to the transport mechanism by AcrB are presented on our blog. We hope that the information provided will be insightful and contribute toward you understanding of the function of AcrB.

2 comments:

Yong Zi30 May 2012 at 13:28
Great introduction and informative video! You mentioned only Gram negative bacteria has AcrB. Do similar proteins exist in Gram positive bacteria?
Ananth31 May 2012 at 14:02
Good question Yong Zi! We had to dig around for a bit to be sure.

I think its worth mentioning here that we have to keep in mind the fact that AcrB is only one of many such molecules involved in drug efflux out of Gram-negative bacteria. There are many other homologues of AcrB such as OprK involved in this process of confering drug resistance to gram-negative bacteria.

When we speak of Gram positive bacteria, the primary distinction that factors into the manner in which drug resistance proteins are expressed, is the structure of the bacteria itself. Gram-positive bacteria have a thick peptidoglycan layer which is non-existent in a Gram-negative bacteria. One such example where this is advantageous is in Vancomycin Resistant Enterocci (VRE). Evolutionarily advantageous, the Gram-positive bacteria simply encodes for further thickening of the peptidoglycan layer. This eventually prevents the vancomycin from reaching its target in the plasma membrane of the Gram-positive bacteria, buried within the peptidoglycan layer. So, in effect, this is a totally different mechanism. Although we must conceed that there has been evidence for multidrug-resistance in Gram-positive bacteria for which the machanism is a little unclear to us at this point. We do hope to find something more on this so that we can address the mechanism involved and to compare if it is any different from the one we described in our blog.

I hope this helps for now!