Beta-lactam antibiotics are used to treat a broad spectrum of Gram-positive and Gram-negative bacteria.
Beta-lactam antibiotics target the penicillin-binding proteins or PBPs – a group of enzymes found attached in the cell membrane, which are involved in the cross-linking of the bacterial cell wall. The beta-lactam ring portion of this group of antibiotics binds to these different PBPs, rendering them unable to perform their role in cell wall synthesis. This then leads to death of the bacterial cell due to osmotic instability or autolysis.
The most effective way for bacteria to counteract these chemicals has been by producing β-lactamases, enzymes that inactivate the drugs by hydrolyzing the β-lactam ring.
β-lactamases are classified under two categories the first one is depending on the biochemical and functional characteristics of the enzyme and secondly based on the molecular structure of the enzyme.
For the functional classification, several criteria are used, including the spectrum of antimicrobial substrate profile, enzyme inhibition profile, hydrolysis rate (Vmax), binding affinity (Km), isoelectric focusing (pI), protein molecular weight, and amino acid composition.
The molecular classification of β-lactamases is based on the nucleotide and amino acid sequences in these enzymes. Four classes of β-lactamases are recognized (A-D), correlating with the functional classification. Classes A, C, and D act by a serine based mechanism, whereas class B or metallo β-lactamases need zinc for their action.
Out of the four classes of β-lactamases class A β-lactamases have been used as versatile scaffolds to create hybrid enzymes, these are referred to as β-lactamase Hybrid Proteins or BHP’s. In this application an exogenous peptide, protein or fragment is inserted at various permissive positions. BHP’s are designed to:
- Create bifunctional proteins.
- To produce and characterize proteins which are otherwise difficult to express.
- Determine the epitope of specific antibodies.
- Generate antibodies against nonimmunogenic epitopes, and
- Better understand the structure/functional relationship of proteins.
The two class A β-lactamases that have been extensively used as model proteins to insert exogenous polypeptides are:
- BlaP from Bacillus licheniformis 749/C and
TEM-1 is the most commonly found β-lactamase in gram-negative bacteria. These two enzymes are, secreted by bacteria in the external environment and expressed in the periplasmic space to inactivate β-lactam antibiotics. About 20% – 40% of the amino acids are, identical and homologous between these two serine-active β-lactamases.
In addition to the above-mentioned applications there are other proposed applications that can be envisaged are:
- BHPs could be used as biosensors and in affinity chromatography, drug screening, and drug targeting.
- They are also of special interest to better understand more fundamental aspects of protein evolution and structure/function relationships.
Beta-lactam Antibiotics: From Antibiosis to Resistance and Bacteriology – Kok-Fai Kong, Lisa Schneper and Kalai Mathee
Class A β-Lactamases as Versatile Scaffolds to Create Hybrid Enzymes: Applications from Basic Research to Medicine – Céline Huynen, Patrice Filée, André Matagne, Moreno Galleni, and Mireille Dumoulin