Linked Life Data

15100473

Source:http://linkedlifedata.com/resource/pubmed/id/15100473

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
2004-4-21
pubmed:abstractText
In a preliminary communication we described a top-down approach to the determination of chemical cross-link location in proteins using Fourier transform mass spectrometry (FT-MS). We have since extended the approach to use a series of homobifunctional cross-linkers with the same reactive functional groups, but different cross-linker arm lengths. Correlating cross-linking data across a series of related linkers allows the distance constraint derived from a cross-link between two reactive side chains to be determined more accurately and increases the confidence in the assignment of the cross-links. In ubiquitin, there are seven lysines with primary amino groups and the amino terminus. Disuccinimidyl suberate (DSS, cross-linker arm length = 11.4 A), disuccinimidyl glutarate (DSG, cross-linker arm length = 7.5 A) and disuccinimidyl tartrate (DST, cross- linker arm length = 5.8 A) are homobifunctional cross-linking reagents that react specifically with primary amines. Using tandem mass spectrometry (MS/MS) on the singly, internally cross-linked precursor ion of ubiquitin, we found cross-links with DSS and DSG between the amino terminus and Lys 6, between Lys 6 and Lys 11, and between Lys 63 and Lys 48. Using disuccinimidyl tartrate (DST), the shortest cross-linker in the series, only the cross-links between the amino terminus and Lys 6, and between Lys 6 and Lys 11 were observed. The observed cross-links are consistent with the crystal structure of ubiquitin, if the lysine side chains and the amino terminus are assumed to have considerable flexibility. In a separate study, we probed the reactivity of the primary amino groups in ubiquitin using the amino acetylating reagent, N-hydroxy succinimidyl acetate (NHSAc), and a top-down approach to localize the acetylated lysine residues. The reactivity order obtained in that study (M1 approximate, equals K6 approximate, equals K48 approximate, equals K63) > K33 > K11 > (K27, K29), shows that the cross-link first formed in ubiquitin by reaction with DSS and DSG occurs between the most reactive residues.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
1469-0667
pubmed:author
pubmed:issnType
Print
pubmed:volume
9
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
623-31
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2003
pubmed:articleTitle
A top-down approach to protein structure studies using chemical cross-linking and Fourier transform mass spectrometry.
pubmed:affiliation
Sandia National Laboratories, PO Box 969, Livermore, CA 94551-0969, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, Non-U.S. Gov't