Chavan A J, Kim H, Haley B E, Watt D S
Department of Chemistry, University of Kentucky, Lexington 40506.
Bioconjug Chem. 1990 Sep-Oct;1(5):337-44. doi: 10.1021/bc00005a007.
A GTP photoaffinity probe (125I-APTG) was developed that incorporated an [125I]-N-(4-azidophenyl)-2-amino-3-(4-hydroxy-3-iodophenyl)propionamide group at the gamma-position of GTP through a phosphonamide linkage. A combination of saturation and GTP protection studies (90% protection at 25 microM GTP with an apparent Kd of 5 microM) validated the use of this new probe as a satisfactory GTP mimic. This probe offered the advantage of possessing an 125I radiolabel external to the GTP moiety, in contrast to the previously reported [gamma 32P]-8-N3GTP that possessed an internal 32P radiolabel. This novel feature accommodated the purification of photolabeled peptides using a combination of ion-exclusion, gel filtration, and HPLC techniques. [125I]APTG was used to identify a peptide (beta:65-79) in the exchangeable GTP-binding domain of the beta-subunit of tubulin.
