Design of new benzoxazole-2-thione-derived inhibitors of Streptococcus pneumoniae hyaluronan lyase: structure of a complex with a 2-phenylindole.

The bacterial hyaluronan lyases (Hyals) that degrade hyaluronan, an important component of the extracellular matrix, are involved in microbial spread. Inhibitors of these enzymes are essential in investigation of the role of hyaluronan and Hyal in bacterial infections and constitute a new class of antibiotics against Hyal-producing bacteria. Recently, we identified 1,3-diacetylbenzimidazole-2-thione and related molecules as inhibitors of streptococcal Hyal.

Structural studies and mechanism of Saccharomyces cerevisiae dolichyl-phosphate-mannose synthase: insights into the initial step of synthesis of dolichyl-phosphate-linked oligosaccharide chains in membranes of endoplasmic reticulum.

Dolichyl-phosphate-mannose (Dol-P-Man) synthase catalyzes the reversible formation of a key intermediate that is involved as a mannosyl donor in at least three different pathways for the synthesis of glycoconjugates important for eukaryotic development and viability. The enzyme is found associated with membranes of the endoplasmic reticulum (ER), where it transfers mannose from the water soluble cytoplasmic donor, guanosine 5'-diphosphate (GDP)-Man, to the membrane-bound, extremely hydrophobic, and long-chain polyisoprenoid acceptor, dolichyl-phosphate (Dol-P). The enzyme from Saccharomyces cerevisiae has been utilized to investigate the structure and activity of the protein and interactions of the enzyme with Dol-P and synthetic Dol-P analogs containing fluorescent probes.

Expression and purification of Streptococcus pneumoniae hyaluronate lyase from Escherichia coli.

Pneumococcal hyaluronate lyase enzyme breaks down hyaluronan of the extracellular matrix of tissues and possibly contributes to the invasion of host tissue and to the penetration of host defenses by this bacterial pathogen. In light of the emergence of increasing numbers of antibiotic-resistant strains, the understanding of the mechanism of action of hyaluronate lyase enzyme may lead to a better understanding of interactions between a host and bacterial pathogens and may contribute to more efficient treatment of bacterial infections. The native Streptococcus pneumoniae hyaluronate lyase enzyme has a molecular mass of 107 kDa but undergoes conversion to smaller enzymatically active forms.

Crystallization and preliminary X-ray analysis of Streptococcus pneumoniae hyaluronate lyase.

A fully active 83-kDa truncated form of recombinant hyaluronate lyase from Streptococcus pneumoniae was crystallized by the hanging drop vapor diffusion method using ammonium sulfate as a precipitating agent. Crystals grew at room temperature using a variety of buffers with pH around 6. The crystals diffract X-rays beyond 2.