CLONING AND SEQUENCING OF ATP SULFURYLASE FROM PENICILLIUM-CHRYSOGENUM - IDENTIFICATION OF A LIKELY ALLOSTERIC DOMAIN

Fungal (Penicillium chrysogenum) and yeast (Saccharomyces cerevisiae) ATP sulfurylases were shown to have very similar kinetic and chemical properties except that the fungal enzyme (a) contains a highly reactiv e Cys residue (SH-1) whose modification results in sigmoidal velocity curves (Renosto, F, Martin, R. L., and Segel, I. H. (1987) J. Biol. Ch em. 262, 16279-16288) and (b) is allosterically inhibited by 3'-phosph oadenosine 5'phosphosulfate (PAPS), while the yeast enzyme displays ne ither of these properties. The fungal enzyme subunit (64.3 kDa, 572 am ino acids) is also larger than the yeast enzyme subunit (59.3 kDa, 521 amino acids). To correlate the unique allosteric properties of the fu ngal enzyme with specific structural features, we cloned and sequenced the ATP sulfurylase gene taps) from P. chrysogenum. The yeast and fun gal enzymes are homologous over the first 400 amino acids and contain two regions high in basic residues which are conserved in sulfurylases from Arabidopsis and the Riftia pachyptila (hydrothermal vent tube wo rm) chemolithotrophic symbiont, These regions may participate in formi ng the binding sites for MgATP(2-) and SO42-. The fungal enzyme has no significant sequence homology to the yeast enzyme in the C-terminal 1 72 amino acids. This C-terminal region contains SH-1 (Cys-508) and has homology to MET14 (S. cerevisiae), CYSC (E. coli), and NODQ (Rhizobiu m meliloti), i.e. adenosine 5'-phosphosulfate (APS) kinase. The cumula tive results suggest that (a) the allosteric PAPS binding site of P. c hrysogenum ATP sulfurylase is located in the C-terminal domain of the protein and (b) that this domain may have evolved from APS kinase. In spite of the homology, this C-terminal region does not account for the APS kinase activity of P. chrysogenum, Fungal ATP sulfurylase has no significant homology to (or regulatory properties in common with) CYSD or CYSN, proteins reported to comprise E. coli ATP sulfurylase (Leyh, T., Vogt, T. F, and Sue, Y. (1992) J. Biol. Chem. 267, 10405-10410).