Hydrolytically degradable thiol-ene hydrogels for protein release.

A new degradable PEG-diester-dinorbornene/PEG-triester-trithiol hydrogel was evaluated for protein release. The hydrogel polymerized rapidly with seconds of UV irradiation and subsequently hydrolytically degraded in aqueous buffer over the course of approximately 3 weeks. Further, the hydrogel enabled the encapsulation and release of a model protein, bovine serum albumin (BSA), over 7 days with ~ 90% released at 48 h.

High-level expression of Escherichia coli and Bacillus subtilis thymidylate synthases.

Procedures are described for the preparation of highly purified thymidylate synthases from Escherichia coli and Bacillus subtilis. The yields in each case are quite high with about 350 mg of pure protein obtained from 1 liter of cells. Basically all that is required to obtain pure enzyme is an induction step from a high-expression vector, followed by a DE-52 column elution.

Nucleosides as a carbon source in Bacillus subtilis: characterization of the drm-pupG operon.

In Bacillus subtilis, nucleosides are readily taken up from the growth medium and metabolized. The key enzymes in nucleoside catabolism are nucleoside phosphorylases, phosphopentomutase, and deoxyriboaldolase. The characterization of two closely linked loci, drm and pupG, which encode phosphopentomutase (Drm) and guanosine (inosine) phosphorylase (PupG), respectively, is reported here.

Crystal structure of thymidylate synthase A from Bacillus subtilis.

Thymidylate synthase (TS) converts dUMP to dTMP by reductive methylation, where 5,10-methylenetetrahydrofolate is the source of both the methylene group and reducing equivalents. The mechanism of this reaction has been extensively studied, mainly using the enzyme from Escherichia coli. Bacillus subtilis contains two genes for TSs, ThyA and ThyB.