LACTB E.coli
b-Lactamase, EC 3.5.2.6, TEM-1.
Greater than 95.0% as determined by SDS-PAGE.
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Description
Recombinant E.coli Beta-Lactamase produced in E.Coli is a single, non-glycosylated polypeptide chain containing 264 amino acids and having a molecular mass of approximately 28.9 kDa.
Beta Lactamase is purified by proprietary chromatographic techniques.
Product Specs
Beta-lactamase, an enzyme classified under EC 3.5.2.6, is produced by certain bacteria and confers resistance to beta-lactam antibiotics. This class of antibiotics, including penicillins, cephalosporins, cephamycins, and carbapenems, share a characteristic four-atom ring structure called a beta-lactam. Beta-lactamase enzymes hydrolyze this ring, effectively neutralizing the antibiotic's antibacterial properties.
Recombinant E. coli Beta-Lactamase, expressed in E. coli, is a single, non-glycosylated polypeptide chain composed of 264 amino acids, with an approximate molecular weight of 28.9 kDa. The purification process involves proprietary chromatographic methods.
Lyophilized from a concentrated solution of 100mM Tris buffer at pH 7.0.
For reconstitution, it is recommended to dissolve the lyophilized Beta-Lactamase in sterile 18 MΩ-cm H₂O to achieve a concentration of 100 µg/ml. This solution can be further diluted in other aqueous solutions as needed. Ensure that the Beta-Lactamase is used within a pH range of 7.0-8.0 and at a temperature not exceeding 45°C.
Greater than 95.0% purity as determined by SDS-PAGE analysis.
One unit of enzyme activity is defined as the amount that hydrolyzes 1.0 µmole of benzyl penicillin per minute at pH 7.0 and 25°C, in the presence of EDTA.
b-Lactamase, EC 3.5.2.6, TEM-1.
Product Science Overview
Beta-lactamases are enzymes produced by bacteria that provide resistance to beta-lactam antibiotics such as penicillins and cephalosporins. These enzymes achieve this by breaking the beta-lactam ring, a crucial component of these antibiotics, thereby neutralizing their antibacterial properties . The recombinant production of beta-lactamase in Escherichia coli (E. coli) has become a significant area of research due to its clinical implications in antibiotic resistance.
Recombinant DNA technology involves the insertion of a gene of interest into a host organism to produce a desired protein. In the case of beta-lactamase, the gene encoding this enzyme is inserted into E. coli, a commonly used host due to its high expression capability and ease of genetic manipulation . The process typically involves the use of cloning vectors such as pET-28a, which allows for the high-level expression of the target protein under the control of an inducible promoter .
The expression of recombinant beta-lactamase in E. coli involves several steps:
- Cloning: The gene encoding beta-lactamase is cloned into a suitable vector, such as pET-28a, which contains elements necessary for the expression of the gene in E. coli .
- Transformation: The recombinant vector is introduced into E. coli cells, typically using a strain like BL21 (DE3), which is engineered to express high levels of the target protein .
- Induction: The expression of beta-lactamase is induced using a chemical such as IPTG (isopropyl-β-D-thiogalactopyranoside), which activates the promoter controlling the gene .
- Purification: The recombinant protein is purified using affinity chromatography, often involving a His-tag that facilitates binding to a nickel column .
The industrial production of recombinant beta-lactamase involves scaling up the expression and purification processes to produce large quantities of the enzyme. This requires optimization of various parameters, including the concentration of the inducer, culture temperature, and induction time . Additionally, the purification process must be efficient to ensure high yields of the target protein with minimal contaminants .
Beta-lactamases have a significant clinical impact as they mediate resistance to beta-lactam antibiotics, which are widely used to treat bacterial infections . The recombinant production of these enzymes in E. coli allows for detailed studies of their structure and function, which can inform the development of new antibiotics and strategies to combat antibiotic resistance .
