Four molecular approaches to determining the types of Candida albicans strains were compared. The strains used were those whose repeated DNA (ribosomal and mitochondrial) EcoRI restriction fragment length polymorphisms (RFLP) were determined by Stevens et al. (D. A.
In Escherichia coli, the phosphorylation and dephosphorylation of isocitrate dehydrogenase (IDH) are catalyzed by a bifunctional protein kinase/phosphatase. We have determined the nucleotide sequence of aceK, the gene encoding IDH kinase/phosphatase. This gene consists of a single open reading frame of 1,734 base pairs preceded by a Shine-Dalgarno ribosome-binding site. Examination of the deduced aminQ acid sequence of IDH kinase/phosphatase revealed sequences which are similar to the consensus sequence for ATP-binding sites. This protein did not, however, exhibit the extensive sequence homologies which are typical of other protein kinases. Multiple copies of the REP family of repetitive extragenic elements were found within the intergenic region between aceA (encoding isocitrate lyase) and aceK. These elements have the potential for combining to form an exceptionally stable stem-loop structure (AG = -54 kcal/mol [ca. -226 kJ/moll) in the miRNA. This structure, which masks the ribosome-binding site and start codon for aceK, may contribute to the downshift in expression observed between aceA and aceK. Another potential stem-loop structure (AG = -29 kcal/mol [ca. 121 kJ/mol]), unrelated to the REP sequences, was found within aceK.For Escherichia coli, adaptation to growth on acetate requires the induction of the enzymes of the glyoxylate bypass, isocitrate lyase and malate synthase (26,27). This pathway allows the net synthesis of cellular constituents from acetate because it bypasses the CO2-producing steps of the Krebs cycle. Once induced, this pathway is regulated, at least in part, through the phosphorylation of isocitrate dehydrogenase (IDH), the Krebs cycle enzyme which competes with isocitrate lyase (3,17,18,21,54). This phosphorylation site has been identified as serine 113 (5, 51). During growth on acetate, ca. 70% of the IDH is maintained in the inactive, phosphorylated form (4, 30, 31), forcing isocitrate through the glyoxylate bypass (32, 41). Mutant strains which fail to express IDH kinase/phosphatase are usually unable to grow on acetate, indicating that the phosphorylation of IDH is essential for growth on this carbon source (31).The phosphorylation and dephosphorylation of IDH are catalyzed by a bifunctional enzyme, IDH kinase/phosphatase. This protein is expressed from a single gene, aceK (28), and yields a single band on sodium dodecyl sulfate electrophoresis (28, 29). These observatioihs support the proposal that IDH kinase and IDH phosphatase reside on the same polypeptide. The physical association of opposing regulatory activities is not unique to IDH kinase/phosphatase. Other bifunctional regulatory proteins have been identified in mammals (14), plants (6), and microorganisms (7,16,42).In addition to regulation mediated by the phosphorylation of IDH, the glyoxylate bypass is regulated at the level of gene expression. Using genetic techniques, Maloy and Nunn demonstrated that the genes which encode isocitrate lyase (aceA) and malate synthase (aceB) are expressed from the same operon and that a...
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