Yeast methionine aminopeptidase type 1 is ribosome‐associated and requires its N‐terminal zinc finger domain for normal function in vivo*

Abstract

Methionine aminopeptidase type 1 (MetAP1) cotranslationally removes N-terminal methionine from nascent polypeptides, when the second residue in the primary structure is small and uncharged. Eukaryotic MetAP1 has an N-terminal zinc finger domain not found in prokaryotic MetAPs. We hypothesized that the zinc finger domain mediates the association of MetAP1 with the ribosomes and have reported genetic evidence that it is important for the normal function of MetAP1 in vivo. In this study, the intracellular role of the zinc finger domain in yeast MetAP1 function was examined. Wild-type MetAP1 expressed in a yeast map1 null strain removed 100% of N-terminal methionine from a reporter protein, while zinc finger mutants removed only 31-35%. Ribosome profiles of map1 null expressing wild-type MetAP1 or one of three zinc finger mutants were compared. Wild-type MetAP1 was found to be an 80S translational complex-associated protein that primarily associates with the 60S subunit. Deletion of the zinc finger domain did not significantly alter the ribosome profile distribution of MetAP1. In contrast, single point mutations in the first or second zinc finger motif disrupted association of MetAP1 with the 60S subunit and the 80S translational complex. Together, these results indicate that the zinc finger domain is essential for the normal processing function of MetAP1 in vivo and suggest that it may be important for the proper functional alignment of MetAP1 on the ribosomes.

Characterization of Two New Aminopeptidases in Escherichia coli

“…The exact function of the N-terminal domains of YpdF and its homologs remains elusive. The N-terminal domains of methionyl aminopeptidases in S. cerevisiae harbor zinc finger motifs and short stretches of basic amino acids, which are suggested to be responsible for binding to the ribosome (20). However, in YpdF and its homologs, we did not observe a similar compositional bias in the N termini.…”

Environmental Regulation of Prions in Yeast

“…We analyzed the RNA regulatory potential for Map1p (a methionine aminopeptidase), for which the catalytic domain (peptidase) may be well separated from the RNAbinding sites. The protein contains two Zn-finger domains, which are essential for the normal processing function of MetAP in vivo [33] and were thought to provide interaction with the ribosome [35]. However, Zn-finger domains have been widely seen to mediate protein-DNA or protein-RNA interactions [34] and hence, they may act as RNA-binding motifs in Map1p as well.…”

“…Map1p is a methionine aminopeptidase (MetAP) that catalyzes the co-translational removal of Nterminal methionine from nascent polypeptides, and it is functionally redundant with Map2p [31,32]. Notably, Map1p contains two zinc-finger motifs, one CCCC-type and the other of the CCHHtype [33], which occur in DNA-binding proteins and in some RBPs [34] -however these domains were not thought to provide selective RNA-binding but rather to confer interaction of Map1p with the ribosome [35]. Yeast cells bearing a plasmid with MAP1 under the control of galactose inducible promoter, and control cells containing an empty plasmid, were grown to mid-log phase and expression was induced with 2% galactose for 1.5 hours.…”

An Unusual Peptide Deformylase Features in the Human Mitochondrial N-terminal Methionine Excision Pathway

“…Tel. : 33-1-69-82-36removed from the mature form and is essential for cell function, possibly allowing interaction with ribosomes (18). PDFs constitute a growing family of hydrolytic enzymes related to the thermolysin-metzincin HEXXH motif-containing family of metalloproteases (6).…”