Bacterial recombination system

Another obvious question is why transfection requires intact genomes, although half molecules, at least overlapping halves (Havender and Trautner, 1972), could be thought to be substrates for primary recombination in transfection. The explanation may be found in the observation, that it is the bacterial recombination system which mediates primary recombination. Recombination in B. subtilis transformation is known to occur via single-... 

One of the best-studied carrier molecules is produced as a primary excretory constituent of the adult male mouse, known from its consistent high concentration as the major urinary protein (MUP). The basic 3-D structure of the protein was initially obtained from a monoclinic crystal of recombinant protein (MUP-I), constructed by induction in a bacterial expression system and purified to homogeneity (Kuser, 1990). A wild type version of MUP finally yielded to NMR analysis a clone of the r-isoform (162 residues) was labelled and compared with the crystal-structure (Lucke et al., 1990). Two views of the molecule... 

Mergulhao, F.J.M., Monteiro, G.A., Cabral, J.M.S., and Taipa, M.A. 2004. Design of bacterial vector systems for the production of recombinant proteins in Escherichia coli. Journal of Microbiology and Biotechnology. 14, 1-14. 

The success of Chapman and co-workers in expression of flavocytochrome 2 in E. coli [23] is encouraging in its impUcations for future expression of flavoproteins in this host because, in their experience both the flavin and heme groups are incorporated into the recombinant protein. Moreover, the bacterial expression system produces the protein 500-1000 fold more efficiently than the yeast from which it was cloned. The enzyme produced in E. coli, however, lacks the first five amino acid residues at its amino terminus, a result which presumably reflects subtle differences in protein synthesis between the two organisms. 

Pharmacology Anakinra is a recombinant, nonglycosylated form of the human interleukin-1 receptor antagonist (IL-1 Ra). Anakinra differs from native human IL-IRa in that it has a single methionine residue at its amino terminus. It is produced by recombinant DNA technology using an Escherichia coli bacterial expression system. 

A number of B. pertussis (polypeptide) antigens have been expressed in E. coli and other recombinant systems. Several of these are being evaluated as potential subunit vaccines, including B. pertussis surface antigen, adhesion molecules and pertussis toxin. Pertussis toxin has been shown to protect mice from both aerosol and intracerebral challenge with virulent B. pertussis. The bacterial proteins that mediate surface adhesion protect mice from aerosol but not intracerebral challenge. Future pertussis subunit vaccines may well contain a combination of two or more pathogen-derived polypeptides. 

Re combinational DNA repair of a circular bacterial chromosome, while essential, sometimes generates deleterious byproducts. The resolution of a Holliday junction at a replication fork by a nuclease such as RuvC, followed by completion of replication, can give rise to one of two products the usual two monomeric chromosomes or a contiguous dimeric chromosome (Fig. 25-41). In the latter case, the covalently linked chromosomes cannot be segregated to daughter cells at cell division and the dividing cells become stuck. A specialized site-specific recombination system in E. coli, the XerCD system, converts the dimeric chromosomes to monomeric chromosomes so that cell division can proceed. The reaction is a site-specific deletion reaction (Fig. 25-39b). This is another example of the close coordination between DNA recombination processes and other aspects of DNA metabolism. 

C. C. Boesen, S. A. Motyka, A. Patamawenu, and P. D. Sun, Development of a recombinant bacterial expression system for tbe active form of a human transforming growtb factor P type II receptor ligand binding domain, Prot. Expr. Purif. 2000, 20, 98-104. 

Transgenic bacterial biosensors. Systems such as the Microtox assay detailed earlier use the marine species Vibrio fischeri as the sensor. Because it uses a marine bacterium, Microtox must be conducted in saline solution, which is ecologically irrelevant for most soils. Because no naturally luminescent soil bacteria are known that could be used as an alternative, one solution is to fuse the genes responsible for bioluminescence into soil-dwelling strains using recombinant technology (Paton et al., 1997). Two approaches can be used ... 

A research team has used recombinant DNA methods to clone and express an unusual protein from a parasitic worm. This protein has a sequence of histidine residues at its N-terminal end and, for laboratory purposes, is produced in bacterial cells. How might this protein be purified from a mixture of other proteins in the bacterial expression system ... 

The natural protein from the worm is known to bind long-chain fatty acids, but the recombinant protein from the laboratory was initially found to have less binding affinity than expected. One possibility was that the recombinant protein site was already occupied by fatty acids from the bacterial cell system. How might such contaminants be removed ... 

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