Evidence that two zinc fingers in the methionine aminopeptidase from Saccharomyces cerevisiae are important for normal growth

282

232

We previously characterized a methionine aminopeptidase (EC 3.4.11.18; Met-AP1; also called peptidase M) in Saccharomyces cerevisiae, which differs from its prokaryotic homologues in that it (i) In all living cells, protein synthesis is initiated with either methionine (in the cytosol of eukaryotes), or formylmethionine (in prokaryotes, mitochondria, and chloroplasts). Where formylmethionine is used, the formyl group is usually removed cotranslationally by a deformylase, leaving methionine bearing a free NH2 group. The methionine NH2 is then removed by one or more methionine aminopeptidases (Met-APs), if the penultimate residue is small and uncharged-e.g

Section: Ggccacagtatcgcaccatatcgtatccacggcggtaaatccgttccc G H S I a Psupporting

confidence: 75%

Amino-terminal protein processing in Saccharomyces cerevisiae is an essential function that requires two distinct methionine aminopeptidases.

Chang²

1995

282

232

“…Experimental results imply that the inhibition of MetAP peptidase activity disrupts the intricate balance between positive and negative regulators of angiogenesis (29,30). Although these compounds have been used primarily in cancer therapy, they also may be effective in noncancerous diseases of aberrant vascularization (29,30) and the treatment of pathogenic microorganisms, including E. coli (31)(32)(33)(34) and microsporidia in AIDS patients (35).…”

Section: Discussionmentioning

confidence: 99%

The anti-angiogenic agent fumagillin covalently modifies a conserved active-site histidine in the Escherichia coli methionine aminopeptidase

Orville

et al. 1998

142

144

Methionine aminopeptidase (MetAP) exists in two forms (type I and type II), both of which remove the N-terminal methionine from proteins. It previously has been shown that the type II enzyme is the molecular target of fumagillin and ovalicin, two epoxide-containing natural products that inhibit angiogenesis and suppress tumor growth. By using mass spectrometry, N-terminal sequence analysis, and electronic absorption spectroscopy we show that fumagillin and ovalicin covalently modify a conserved histidine residue in the active site of the MetAP from Escherichia coli, a type I enzyme. Because all of the key active site residues are conserved, it is likely that a similar modification occurs in the type II enzymes. This modification, by occluding the active site, may prevent the action of MetAP on proteins or peptides involved in angiogenesis. In addition, the results suggest that these compounds may be effective pharmacological agents against pathogenic and resistant forms of E. coli and other microorganisms.

“…The N-terminal sequence of the yeast MetAP has two putative zinc-finger domains that may allow interactions with nucleic acid (13). The excision of these domains has little effect on the catalytic properties of the enzyme, but the construct expressing this truncated form is significantly less effective in rescuing the slow-growth phenotype of a MetAP mutant as compared with the wild-type MetAP (34). In contrast, the human enzyme lacks the consensus zinc-finger sequences and instead contains the extended runs of basic and acidic residues.…”

Section: Discussionmentioning

confidence: 99%

Eukaryotic methionyl aminopeptidases: two classes of cobalt-dependent enzymes.

Arfin

Kendall

Hall

et al. 1995

235

181

Using partial amino acid sequence data derived from porcine methionyl aminopeptidase (MetAP; methionine aminopeptidase, peptidase M; EC 3.4.11.18), a fulllength clone of the homologous human enzyme has been obtained. The cDNA sequence contains 2569 nt with a single open reading frame corresponding to a protein of 478 amino acids. The C-terminal portion representing the catalytic domain shows limited identity with MetAP sequences from various prokaryotes and yeast, while the N terminus is rich in charged amino acids, including extended strings of basic and acidic residues. These highly polar stretches likely result in the spuriously high observed molecular mass (67 kDa). This cDNA sequence is highly similar to a rat protein, termed p67, which was identified as an inhibitor of phosphorylation of initiation factor eIF2a and was previously predicted to be a metallopeptidase based on limited sequence homology. Model building established that human MetAP (p67) could be readily accommodated into the Escherichia coli MetAP structure and that the Co2+ ligands were fully preserved. However, human MetAP was found to be much more similar to a yeast open reading frame that differed markedly from the previously reported yeast MetAP. A similar partial sequence from Methanothermusfervidus suggests that this p67-like sequence is also found in prokaryotes. These findings suggest that there are two cobalt-dependent MetAP families, presently composed of the prokaryote and yeast sequences (and represented by the E. coli structure) (type I), on the one hand, and by human MetAP, the yeast open reading frame, and the partial prokaryotic sequence (type II), on the other.