; R o o t E n t r y F 4 @ C o m p O b j b W o r d D o c u m e n t O b j e c t P o o l 4 4 F Microsoft Word 6.0 Document MSWordDoc Word.Document.6 ; Oh +' 0 $ H l D h R:WWUSERTEMPLATENORMAL.DOT marcia steinberg marcia steinberg @ qP 4 @ {! e = e | o " " " " " " " L L L L L d n L C x | | | | | | | # ? e V T 4 " | | | | | | " " | x | | | | " | " | 6 > " " " " | | 4 | 9512655 Chang PRIVATE The broad goal of this project is to understand the mechanism of amino terminal processing of eukaryotic proteins. The approach used is the purification, characterization and genetic analysis of two methionine aminopeptidases (MAPs) in Saccharomyces cerevisiae. The first aim is to elucidate the roles of yeast MAP1 in amino terminal processing and cell growth. Yeast MAP 1 contains two zinc fingers and a cobalt dependent catalytic domain. To understand the roles of MAP1 in this processing event, this investigator plans to study the roles of the zinc fingers and cobalt ions in MAP1 function. The results will show whether MAP1 is ribosome associated, and if so, how deletion of zinc fingers affects thi s interaction The P.I. also plans to compare the efficiency of methionine removal of several model proteins in yeast cells that express no MAP1, truncated MAP1 which lacks zinc fingers, and wild type MAP1 and to elucidate the roles of cobalt in MAP1 function by determining the cobalt binding constant and studying the effects of mutations at the cobalt binding sites on cell growth and enzyme activity. The second aim is to elucidate the roles of is MAP2 in N terminal processing and to determine whether it is a bifunctional protein. The P.I. plans to purify and characterize this enzyme, to compare its substrate specificity and other properties with those of MAP1, and to knock out the MAP2 gene alone and both MAPl and MAP2 genes, and characterize the phenotype of the knockout yeast strains. Amino terminal processing is one of the most common protein processing events, which occurs in all living cells and is essential for normal cell growth. Yet, until recently, very little was known about the mechanism by which this important processing event is carried out in eukaryotic cells, and how it is regulated. This study should provide novel insights into the mechanism and regulation of this important event, which in turn may lead to better understanding of the regulation of protein synthesis and the regulation of the functions of myristoylated proteins, including certain oncogene products, viral proteins, and proteins involved in transmembrane signaling and protein secretory vesicular trafficking. %%% This research project is focused on studying the mechanism of the removal of the initial amino acid from the origin of a protein called the N terminus. This is one of the most common systems for modifying newly made proteins. The generality of this phenomenon in proteins from microbes to man indicates its fundamental importance to all living organisms. Yet, relatively little is known about the mechanism by which this initial amino acid is removed, particularly in higher organisms like man, or the manner in wh ich this process is regulated. The Principal Investigator has already made considerable progress in developing the problem. He has found two enzymes which are specific for removing the N terminal methionine (the initial amino acid) from newly synthesized proteins as they are produced in eukaryotic cells. The next immediate goal of his research is to fully characterize these interesting proteins, so called methionine aminopeptidases (MAPs), that are important to every living cell. It is certain that understanding the novel features of yeast MAPs will provide new insights into the mechanism and the regulation of this cellular event that is essential for cell growth. *** @ ....()()))()() Z : phoenix W EH o ; S u m m a r y I n f o r m a t i o n ( 4 @ E 4 @ Microsoft Word 6.0 4
