Characterization of methyltransferase properties of Escherichia coli YabC protein with an enzyme-coupled colorimetric assay.

An enzyme-coupled colorimetric assay for S-adenosylmethionine-dependent methyltransferase (MT) was established to characterize the enzymatic identity of YabC protein from Escherichia coli. Results showed that the MT activity of YabC is able to be effectively detected with this coupling assay system when filamentous cell lysates from an S-adenosylmethionine synthetase mutant, MEW402metK84, were employed as the source of methylation target, whereas no activity exhibited under same conditions from lysates from E. coli normal shape cells.

pYEMF, a pUC18-derived XcmI T-vector for efficient cloning of PCR products.

A 1330-bp DNA sequence with two XcmI cassettes was inserted into pUC18 to construct an efficient XcmI T-vector parent plasmid, pYEMF. The large size of the inserted DNA fragment improved T-vector cleavage efficiency, and guaranteed good separation of the molecular components after restriction digestion. The pYEMF-T-vector generated from parent plasmid pYEMF permits blue/white colony screening; cloning efficiency analysis showed that most white colonies (>75%) were putative transformants which carried the cloning product.

Extraction of erythrocyte enzymes for the preparation of polyhemoglobin-catalase-superoxide dismutase.

In sustained severe ischemia, reperfusion with oxygen carriers may result in ischemia-reperfusion injuries because of the release of damaging oxygen radicals. A nanobiotechnology-based polyhemogloin-calatase-superoxide dismutase can prevent this because the oxygen carrier, polyhemoglobin, is linked to antioxidant enzymes, catalase and superoxide dismutase. However, these antioxidant enzymes come from nonhuman sources and recombinant human enzymes are expensive.