Linked Life Data

15039940

Source:http://linkedlifedata.com/resource/pubmed/id/15039940

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
2004-3-24
pubmed:abstractText
We present a method employing top-down Fourier transform mass spectrometry (FTMS) for the rapid profiling of amino acid side-chain reactivity. The reactivity of side-chain groups can be used to infer residue-specific solvent accessibility and can also be used in the same way as H/D exchange reactions to probe protein structure and interactions. We probed the reactivity of the N-terminal and epsilon-lysine amino groups of ubiquitin by reaction with N-hydroxysuccinimidyl acetate (NHSAc), which specifically acetylates primary amines. Using a hybrid Q-FTMS instrument, we observed several series of multiply acetylated ubiquitin ions that varied with the NHSAc:protein stoichiometry. We isolated and fragmented each member of the series of acetylated ubiquitin ions in the front end of the instrument and measured the fragment ion masses in the FTMS analyzer cell to determine which residue positions were modified. As we increased the NHSAc:protein stoichiometric ratio, identification of the fragments from native protein and protein with successively increasing modification allowed the assignment of the complete order of reactivity of the primary amino groups in ubiquitin (Met 1 approximately Lys 6 approximately Lys 48 approximately Lys 63>Lys 33>Lys 11>Lys 27, Lys 29). These results are in excellent agreement with the reactivity expected from other studies and predicted from the known crystal structure of ubiquitin. The top-down approach eliminates the need for proteolytic digestion, high-performance liquid chromatographic separations and all other chemical steps except the labeling reaction, making it rapid and amenable to automation using small quantities of protein.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
1076-5174
pubmed:author
pubmed:copyrightInfo
Copyright 2004 John Wiley & Sons, Ltd.
pubmed:issnType
Print
pubmed:volume
39
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
322-8
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
A top-down method for the determination of residue-specific solvent accessibility in proteins.
pubmed:affiliation
Sandia National Laboratories, Livermore, California 94551-0969, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S.