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Regulation and biosynthesis of carbapenem antibiotics in bacteria

Key Points

  • Members of the β-lactam family of antibiotics (penicillins, cephalosporins/cephamycins, monobactams, clavams and carbapenems) function by interfering with bacterial cell wall biosynthesis and are in widespread clinical use for the treatment of bacterial infections.

  • Carbapenems are of particular interest because they are often resistant to the many β-lactamase enzymes that are produced by clinically important bacterial pathogens. β-lactamases commonly confer resistance to penicillin and cephalosporin-related antibiotics.

  • Carbapenem-producing organisms include Streptomyces cattleya, which produces thienamycin, and a few Gram-negative bacterial species, including Erwinia carotovora subsp. carotovora, Serratia sp. strain ATCC39006 and Photorhabdus luminescens, which all produce Car (1-carbapen-2-em-3-carboxylic acid).

  • The E. c. carotovora and Serratia ATCC39006 carA–H biosynthetic operons encode genes that are involved in Car biosynthesis (carA–E) and genes that are required for the Car resistance mechanism (carF and carG). Expression of the carA–H operon in E. c. carotovora and Serratia ATCC39006 is regulated by the Hor/Rap transcriptional regulators, respectively. Expression is also regulated by quorum sensing (cell density-dependent) control in both bacteria — although the underlying mechanisms are different. Physiological cues that affect Car production in Erwinia spp. include temperature, oxygen availability and carbon source.

  • The core biosynthetic enzymes are CarA (carbapenam synthetase), CarB (carboxymethylproline synthase) and CarC (carbapenem synthase). CarB (similar to enoyl coenzyme A (CoA) hydratase enzymes) condenses malonyl-CoA and glutamate semialdehyde to form (2S,5S)-carboxymethyl proline (CMP), which is the substrate for the CarA enzyme. CarA catalyses the ATP-dependent formation of (3S,5S)-carbapenam from CMP, resulting in the closure of the β-lactam ring. Finally, CarC acts on the carbapenam product of CarA to introduce a double bond and to produce a stereoinversion, thereby forming the active molecule, carbapenem.

  • Homologues of CarA and CarC (β-lactam synthetase and clavaminate synthase, respectively) function in clavulanic acid biosynthesis in Streptomyces clavuligerus, and the Car and clavulanate biosynthesis pathways have some similarities. Putative homologues of carA (thnM) and carB (thnE) have been found in the thnA–V operon required for thienamycin biosynthesis in S. cattleya, and a biosynthetic pathway for thienamycin has been proposed.

  • An improved understanding of carbapenem biosynthesis might pave the way towards the development of novel carbapenems that have useful chemotherapeutic activities.

Abstract

Carbapenem antibiotics are members of the β-lactam family of antibiotics, the most important class of antibiotics currently in clinical use. They are active against many important Gram-positive and Gram-negative pathogens. One important feature of carbapenem antibiotics is their resistance to several β-lactamases. Thienamycin, isolated from Streptomyces cattleya, was the first carbapenem described. Other well-studied carbapenems were isolated from the Gram-negative bacteria Erwinia carotovora subsp. carotovora, Serratia sp. strain ATCC39006 and Photorhabdus luminescens strain TT01. Here, we review the genetics and biochemistry of carbapenem production in these bacteria. Research into carbapenems could uncover a new repertoire of bioactive molecules and biosynthetic enzymes, and exploiting these novel enzymes could lead to development of new classes of antibiotics with useful chemotherapeutic activities.

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Figure 1: Car antibiotic biosynthesis and resistance gene clusters of three Gram-negative bacteria.
Figure 2: Summary of known regulators of Car biosynthesis in Erwinia species.
Figure 3: Proposed biosynthetic pathway of Car.
Figure 4: CarA and CarC atomic structures.
Figure 5: Thienamycin and the thn gene cluster.
Figure 6: Car production, resistance and quorum sensing in Erwinia carotovora subsp. carotovora and Serratia sp. strain ATCC39006.

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Acknowledgements

The authors would like to thank the BBSRC, Sygen International Plc and NERC for financial support, and would also like to thank Martin Welch and members of the Salmond group for valuable discussions. We dedicate this paper to Barrie Bycroft, on his retirement, for stimulating our collaborative interest in carbapenems.

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Correspondence to George P. C. Salmond.

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DATABASES

Entrez

Erwinia carotovora subsp. atroseptica

Photorhabdus luminescens strain TT01

SwissProt

CarA

CarB

CarC

CarD

CarE

CarF

CarG

CarH

FURTHER INFORMATION

George P. C. Salmond's laboratory

Glossary

FERMENTATION

An industrial process that is used to generate a microbially derived product in a fermenter, in which the growth conditions of the microorganism are tightly controlled and optimized to produce the maximum yield of the required product.

SEMI-SYNTHESIS

This describes a process by which molecules of interest are produced by synthetic chemical modification of starting-point compounds that have been produced by microbial fermentation rather than by total chemical synthesis in vitro.

CRYPTIC GENES

Cryptic genes are present in the genome of the microorganism of interest, but are not generally expressed. Crypticity can be due to mutations in regulatory genes. Alternatively, genes that are expressed might have non-functional protein products owing to mutations in the coding sequences.

INTRINSIC RESISTANCE

Some bacteria are intrinsically resistant to specific antibiotics and have not acquired antibiotic resistance through gene mutation or horizontal gene transfer from another organism. Intrinsic resistance is particularly important for antibiotic-producing organisms, which must have a mechanism for avoiding the inhibitory effects of the antibiotic that they produce.

CONSTITUTIVE EXPRESSION

This describes a constant level of gene expression rather than gene expression that is induced or repressed in response to a change in a physiological cue.

DEREPRESSION

The removal of repression in response to a physiological signal, resulting in an up-regulation of expression of the corresponding target genes.

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Coulthurst, S., Barnard, A. & Salmond, G. Regulation and biosynthesis of carbapenem antibiotics in bacteria. Nat Rev Microbiol 3, 295–306 (2005). https://doi.org/10.1038/nrmicro1128

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