Enzyme activities contributing to hypoxanthine production in Ureaplasma

Several isozymes have been evaluated by other investigators to help characterize both mycoplasmas and acholeplasmas. We have investigated a number of enzymes contributing to hypoxanthine production in Ureaplasma urealyticum, as part of an ongoing effort to identify a comparative profile of isozyme activities in this species. Cells from large volume cultures were collected by centrifugation and lysed by both freeze-thawing and sonication in hypotonic buffer with Triton X-100. Lysate was clarified by centrifugation. Proteins in the cell lysate were separated by polyacrylamide gel electrophoresis, incorporating Triton X-100 in the gel and electrode buffer. Gels were stained to indicate sites of hypoxanthine production from AMP, adenosine, inosine, or adenine, in either phosphate or Tris buffer. The results suggest that adenine deaminase, inosine nucleosidase, and adenosine phosphorylase activities are present in the cell lysate, while adenosine nucleosidase and adenosine deaminase activities are absent. Inosine phosphorylase, AMP nucleosidase and/or 5'-nucleotidase activities may also be present. With the formation of hypoxanthine, the possibility for a salvage pathway exists.