Evidence of a dominant negative mutant of yeast methionine aminopeptidase type 2 in Saccharomyces cerevisiae

Joseph A Vetro, Benjamin Dummitt, William S. Micka, Yie Hwa Chang

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Eukaryotic methionine aminopeptidase type 2 (MetAP2, MetAP2 gene (MAP2)), together with eukaryotic MetAP1, cotranslationally hydrolyzes initiator methionine from nascent polypeptides when the side chain of the second residue is small and uncharged. In this report, we took advantage of the yeast (Saccharomyces cerevisiae) map1 null strain's reliance on MetAP2 activity for the growth and viability to provide evidence of the first dominant negative mutant of eukaryotic MetAP2. Replacement of the conserved His174 with alanine within the C-terminal catalytic domain of yeast MetAP2 eliminated detectable catalytic activity against a peptide substrate in vitro. Overexpression of MetAP2 (H174A) under the strong CPD promoter in a yeast map1 null strain was lethal, whereas overexpression under the weaker GAL1 promoter slightly inhibited map1 null growth. Deletion mutants further revealed that the N-terminal region of MetAP2 (residues 2-57) is essential but not sufficient for MetAP2 (H174A) to fully interfere with map1 null growth. Together, these results indicate that catalytically inactive MetAP2 is a dominant negative mutant that requires its N-terminal region to interfere with wild-type MetAP2 function.

Original languageEnglish (US)
Pages (from-to)656-668
Number of pages13
JournalJournal of Cellular Biochemistry
Volume94
Issue number4
DOIs
StatePublished - Mar 1 2005

Fingerprint

Yeast
Saccharomyces cerevisiae
Yeasts
Growth
Peptides
Alanine
Methionine
Catalytic Domain
Catalyst activity
Genes
methionine aminopeptidase 2
Substrates
In Vitro Techniques

Keywords

  • Angiogenesis
  • Methionine aminopeptidase
  • Protein processing
  • TNP-470
  • p67

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Evidence of a dominant negative mutant of yeast methionine aminopeptidase type 2 in Saccharomyces cerevisiae. / Vetro, Joseph A; Dummitt, Benjamin; Micka, William S.; Chang, Yie Hwa.

In: Journal of Cellular Biochemistry, Vol. 94, No. 4, 01.03.2005, p. 656-668.

Research output: Contribution to journalArticle

Vetro, Joseph A ; Dummitt, Benjamin ; Micka, William S. ; Chang, Yie Hwa. / Evidence of a dominant negative mutant of yeast methionine aminopeptidase type 2 in Saccharomyces cerevisiae. In: Journal of Cellular Biochemistry. 2005 ; Vol. 94, No. 4. pp. 656-668.
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