Linked Life Data

9601050

Source:http://linkedlifedata.com/resource/pubmed/id/9601050

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
21
pubmed:dateCreated
1998-6-12
pubmed:abstractText
p21-activated kinases (PAKs) serve as effector proteins for the GTP-binding proteins Cdc42 and Rac. They are serine/threonine kinases containing the Cdc42/Rac interactive binding (CRIB) motif. The main aim of this study was to define the minimal domain of alphaPAK required for Cdc42/Rac binding. Eight stable PAK fragments of varying lengths, each containing the CRIB motif (residues 75-88), were expressed in Escherichia coli, and their ability to interact with Cdc42 and Rac was assessed using scintillation proximity assays, isothermal titration calorimetry, and fluorescence techniques. The shortest fragments examined (residues 70-94 and 75-94) bound only weakly to either Cdc42 or Rac. A longer fragment starting at residue 75 and ending at residue 105 showed binding to Q61L Rac.GTP with Kd = 1.9 microM. Highest affinity binding (Kd approximately 0.05 microM) was seen with longer fragments ending at residue 118 or 132. A small increase in affinity was seen with those fragments starting at residue 70 rather than residue 75. PAK fragments bound with approximately 3-10-fold higher affinity to Cdc42 than to Rac and bound Q61L variants with 5-10-fold higher affinity than wild type. The dissociation rates of Q61L Rac.mant-GTP and of Q61L Cdc42. mant-GTP from PAK fragment residues 70-132 were measured to be 0.66 and 0.25 min-1, respectively, which are 100-fold lower than dissociation rates for Ras:Ras-effector domains, although their affinities are similar. Calorimetric measurements revealed that binding was associated with a relatively slow heat change. It is suggested that these PAK fragments (in the absence of Cdc42 or Rac) might exist predominantly in an inactive conformation that slowly interconverts with an active conformation and/or a slow conformational change may occur upon binding to Cdc42/Rac. In conclusion, the PAK CRIB motif itself is insufficient for high-affinity binding to Cdc42/Rac, but a 30 amino acid region of PAK (residues 75-105), containing this motif, is sufficient.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
26
pubmed:volume
37
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
7885-91
pubmed:dateRevised
2007-11-19
pubmed:meshHeading
pubmed-meshheading:9601050-Amino Acid Sequence, pubmed-meshheading:9601050-Animals, pubmed-meshheading:9601050-Calorimetry, pubmed-meshheading:9601050-Cell Cycle Proteins, pubmed-meshheading:9601050-GTP-Binding Proteins, pubmed-meshheading:9601050-Genetic Vectors, pubmed-meshheading:9601050-Kinetics, pubmed-meshheading:9601050-Molecular Sequence Data, pubmed-meshheading:9601050-Peptide Fragments, pubmed-meshheading:9601050-Protein Binding, pubmed-meshheading:9601050-Protein Structure, Tertiary, pubmed-meshheading:9601050-Protein-Serine-Threonine Kinases, pubmed-meshheading:9601050-Rats, pubmed-meshheading:9601050-Recombinant Fusion Proteins, pubmed-meshheading:9601050-Scintillation Counting, pubmed-meshheading:9601050-cdc42 GTP-Binding Protein, Saccharomyces cerevisiae, pubmed-meshheading:9601050-p21-Activated Kinases, pubmed-meshheading:9601050-rac GTP-Binding Proteins
pubmed:year
1998
pubmed:articleTitle
Delineation of the Cdc42/Rac-binding domain of p21-activated kinase.
pubmed:affiliation
Exploratory Chemistry Unit, Glaxo Wellcome Medicines Research Centre, Hertfordshire, United Kingdom.
pubmed:publicationType
Journal Article