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Proteins labeled with 32P can be used as sensitive "prime" in blot overlays to detect binding proteins or domains. Small G-protein Ras can bind GTP with extremely high affinity (Kd ∼ 10−11 - 10−12 M) in the presence of Mg2+. We have taken advantage of this property of Ras to develop a vector that expresses proteins of interest such as glutathione S-transferase (GST)/Ras fusion proteins for...
The PAK family of kinases are regulated through interaction with the small GTPases Cdc42 and Rac1, but little is known of the signaling components immediately upstream or downstream of these proteins. We have purified and cloned a new class of Rho-p21 guanine nucleotide exchange factor binding tightly through its N-terminal SH3 domain to a conserved proline-rich PAK sequence with a K d of...
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