Linked Life Data

11463793

Source:http://linkedlifedata.com/resource/pubmed/id/11463793

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
39
pubmed:dateCreated
2001-9-24
pubmed:abstractText
Saccharomyces cerevisiae RNA triphosphatase (Cet1) and RNA guanylyltransferase (Ceg1) interact in vivo and in vitro to form a bifunctional mRNA capping enzyme complex. Here we show that the guanylyltransferase activity of Ceg1 is highly thermolabile in vitro (98% loss of activity after treatment for 10 min at 35 degrees C) and that binding to recombinant Cet1 protein, or a synthetic peptide Cet1(232-265), protects Ceg1 from heat inactivation at physiological temperatures. Candida albicans guanylyltransferase Cgt1 is also thermolabile and is stabilized by binding to Cet1(232-265). In contrast, Schizosaccharomyces pombe and mammalian guanylyltransferases are intrinsically thermostable in vitro and they are unaffected by Cet1(232-265). We show that the requirement for the Ceg1-binding domain of Cet1 for yeast cell growth can be circumvented by overexpression in high gene dosage of a catalytically active mutant lacking the Ceg1-binding site (Cet1(269-549)) provided that Ceg1 is also overexpressed. However, such cells are unable to grow at 37 degrees C. In contrast, cells overexpressing Cet1(269-549) in single copy grow at all temperatures if they express either the S. pombe or mammalian guanylyltransferase in lieu of Ceg1. Thus, the cell growth phenotype correlates with the inherent thermal stability of the guanylyltransferase. We propose that an essential function of the Cet1-Ceg1 interaction is to stabilize Ceg1 guanylyltransferase activity rather than to allosterically regulate its activity. We used protein-affinity chromatography to identify the COOH-terminal segment of Ceg1 (from amino acids 245-459) as an autonomous Cet1-binding domain. Genetic experiments implicate two peptide segments, (287)KPVSLYVW(295) and (337)WQNLKNLEQPLN(348), as likely constituents of the Cet1-binding site on Ceg1.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
28
pubmed:volume
276
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
36116-24
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed-meshheading:11463793-Acid Anhydride Hydrolases, pubmed-meshheading:11463793-Alanine, pubmed-meshheading:11463793-Amino Acid Sequence, pubmed-meshheading:11463793-Animals, pubmed-meshheading:11463793-Binding Sites, pubmed-meshheading:11463793-Candida albicans, pubmed-meshheading:11463793-Catalysis, pubmed-meshheading:11463793-Cell Division, pubmed-meshheading:11463793-Chromatography, Affinity, pubmed-meshheading:11463793-Glutathione Transferase, pubmed-meshheading:11463793-Hot Temperature, pubmed-meshheading:11463793-Molecular Sequence Data, pubmed-meshheading:11463793-Mutagenesis, Site-Directed, pubmed-meshheading:11463793-Mutation, pubmed-meshheading:11463793-Nucleotidyltransferases, pubmed-meshheading:11463793-Phenotype, pubmed-meshheading:11463793-Plasmids, pubmed-meshheading:11463793-Protein Binding, pubmed-meshheading:11463793-Protein Structure, Tertiary, pubmed-meshheading:11463793-RNA, Messenger, pubmed-meshheading:11463793-Recombinant Fusion Proteins, pubmed-meshheading:11463793-Recombinant Proteins, pubmed-meshheading:11463793-Saccharomyces cerevisiae, pubmed-meshheading:11463793-Schizosaccharomyces, pubmed-meshheading:11463793-Sequence Homology, Amino Acid, pubmed-meshheading:11463793-Temperature
pubmed:year
2001
pubmed:articleTitle
An essential function of Saccharomyces cerevisiae RNA triphosphatase Cet1 is to stabilize RNA guanylyltransferase Ceg1 against thermal inactivation.
pubmed:affiliation
Molecular Biology Program, Sloan-Kettering Institute, New York, New York 10021, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.