Linked Life Data

2036409

Source:http://linkedlifedata.com/resource/pubmed/id/2036409

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
22
pubmed:dateCreated
1991-7-2
pubmed:abstractText
A method is presented for the incorporation of nonnatural amino acids into proteins during in vitro cell-free translation. A combination of chemical synthesis and run-off transcription was employed to prepare a semisynthetic, nonhypermodified tRNA(Gly) nonsense suppressor acylated with L-3-[125I]iodotyrosine. The presence of this synthetic tRNA during in vitro translation of mRNA containing a nonsense suppression site (e.g., a UAG termination codon) results in the incorporation of the nonnatural amino acid L-3-iodotyrosine into the polypeptide exclusively at the position corresponding to that site. Incorporation of the nonnatural amino acid L-3-[125I]iodotyrosine into the model polypeptide was assessed by quantitative and unambiguous determination of suppression efficiency, read-through, and site specificity of incorporation. Minor modifications of the method employed in this initial experiment also allow the rapid analysis of unlabeled acylated tRNA analogues. Under optimum conditions, the unlabeled amino acid L-3-iodotyrosine was found to be incorporated with a suppression efficiency of 65%. Other nonnatural residues, including N-methylphenylalanine, D-phenylalanine, and phenyllactic acid, were tested in the assay under these same conditions. Suppression efficiencies for this series ranged from 0 to 72% depending on the structure of the residue incorporated. Several other aspects of this methodology, such as tRNA structure and context effects, are briefly discussed.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
4
pubmed:volume
30
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
5411-21
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1991
pubmed:articleTitle
Site-specific incorporation of nonnatural residues during in vitro protein biosynthesis with semisynthetic aminoacyl-tRNAs.
pubmed:affiliation
Department of Biological Chemistry, University of California, Irvine 92717.
pubmed:publicationType
Journal Article, Comparative Study, In Vitro, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't