Linked Life Data

7989377

Source:http://linkedlifedata.com/resource/pubmed/id/7989377

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
50
pubmed:dateCreated
1995-1-12
pubmed:abstractText
We have searched for irreversible ligands which target the guanine nucleotide binding pocket of G protein alpha-subunits by testing the ability of periodate-oxidized 2',3'-dialdehyde guanine nucleotide analogues of GTP (oGTP) and GTP gamma S (oGTP gamma S) to bind to the recombinant alpha-subunit of the stimulatory G protein, rGs alpha-s. oGTP and oGTP gamma S bind to rGs alpha-s in a quasi-irreversible manner via formation of a Schiff's base, which can be reduced with borhydrid resulting in covalent incorporation of [alpha-32P]oGTP and [35S]oGTP gamma S into rGs alpha-s. When bound to rGs alpha-s, oGTP is hydrolyzed and traps the protein in the inactive conformation, while oGTP gamma S persistently activates rGs alpha. Thus, oGTP and oGTP gamma S act as irreversible G protein antagonist and agonist, respectively, and represent a pair of nucleotide analogues suitable as functional and structural tools. Cleavage of covalently labeled rGs alpha-s with cyanogen bromide generates several labeled fragments. Labeled fragments were assigned to the G1 and G4 region of the guanine nucleotide binding pocket using sequence-specific antisera. An additional, labeled fragment was identified by amino-terminal sequencing and corresponded to the helix alpha A in the recently determined crystal structure of the transducin alpha-subunit (Noel, J. P., Hamm, H. E., and Sigler, P. B. (1993) Nature 366, 654-663). In the oGDP-liganded conformation, incorporation occurs predominantly into the G1-fragment, while [35S]oGTP gamma S labels the additional fragments to a similar extent indicating tight packing around the guanine nucleotide binding pocket in the active conformation. Furthermore, rGs alpha-s contains a single acid cleavable bond (Asp317-Pro318), such that formic acid releases a carboxyl-terminal fragment from [alpha-32P]oGTP- and [35S]oGTP gamma S-liganded rGs alpha-s. This fragment contains a single lysine residue (Lys324) which is only labeled by [35S]oGTP gamma S. Lys324 is unique to Gs alpha and lies within its effector binding region. Hence, during the switch from the inactive to the active state, this region undergoes a major conformational change that moves it closer to the nucleotide binding pocket.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
16
pubmed:volume
269
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
32008-15
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed-meshheading:7989377-Adenylate Cyclase, pubmed-meshheading:7989377-Amino Acid Sequence, pubmed-meshheading:7989377-Binding Sites, pubmed-meshheading:7989377-Cerebral Cortex, pubmed-meshheading:7989377-DNA Mutational Analysis, pubmed-meshheading:7989377-GTP-Binding Proteins, pubmed-meshheading:7989377-Guanine Nucleotides, pubmed-meshheading:7989377-Guanosine 5'-O-(3-Thiotriphosphate), pubmed-meshheading:7989377-Guanosine Triphosphate, pubmed-meshheading:7989377-Humans, pubmed-meshheading:7989377-Ligands, pubmed-meshheading:7989377-Molecular Sequence Data, pubmed-meshheading:7989377-Oxidation-Reduction, pubmed-meshheading:7989377-Peptide Fragments, pubmed-meshheading:7989377-Protein Conformation, pubmed-meshheading:7989377-Receptors, Purinergic P1, pubmed-meshheading:7989377-Recombinant Proteins, pubmed-meshheading:7989377-Signal Transduction, pubmed-meshheading:7989377-Structure-Activity Relationship
pubmed:year
1994
pubmed:articleTitle
Structural and functional characterization of the interaction between 2',3'-dialdehyde guanine nucleotide analogues and the stimulatory G protein alpha-subunit.
pubmed:affiliation
Institute of Pharmacology, University of Vienna, Austria.
pubmed:publicationType
Journal Article, In Vitro, Research Support, Non-U.S. Gov't