Research projects

The main goal of the project has been to elucidate the possible moonlighting rol of the enzyme N-acetyl glucosamine (NAGA) kinase from the non-conventional yeast
Yarrowia lipolytica.

The moonlighting function of NAGA kinase (YlNag5) was suggested by the changes observed in the expression pattern of genes encoding enzymes from the NAGA catabolic pathway (NAG genes) in a mutant lacking YlNAG5. The obtainment of a double mutant Ylnag5 Ylngt1 (lacking the NAGA transporter) showed that the alteration of the expression of the NAG genes was not due to an internal accumulation of NAGA.

To ascertain if the enzymatic and the regulatory roles of YlNag5 could be separated, heterologous NAGA kinases and tagged versions of YlNAG5 were expressed in a Ylnag5
mutant. The results indicated that both functions could be separated. To obtain further evidence, we generated a library of variants of random mutagenized YlNAG5. Our results are
consistent with a moonlighting function of YlNag5. In addition, in these experiments we demonstrated the NAGA kinase function of a protein of the plant pathogen Magnaporthe
oryzae.

We studied the interaction of YlNag5 with other proteins and identified one with 84 %similarity with the ribosomal protein Urp2 from Saccharomyces cerevisiae. RNA seq experiments with a Ylnag5 mutant showed changes in the expression of genes encoding proteins related with the cell wall or containing a GPI anchor.

A fragment of 121 bp of the promoter region of YlNAG5 is sufficient to direct its transcription conserving all the observed regulatory properties. The sequences cccyrka and ctkrth were found in the NAG genes suggesting them as candidates to bind common regulatory elements.

We noticed the appearance of a mutation that alters the expression pattern of the NAG genes and have shown that the genes RON1 and NGS1, regulators of their expression in other yeast species, are not involved in the phenomenon.

By expressing the YlNAG1 gene encoding the glucosamine-6-P deaminase in S. cerevisiae we obtained a strain able to grow in glucosamine, solving an unexplained metabolic problem in this model yeast.