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A photoactive phosphonamide derivative of GTP for the identification of the GTP-binding domain in beta-tubulin

Journal Article · · Bioconjugate Chemistry; (United States)
DOI:https://doi.org/10.1021/bc00005a007· OSTI ID:5272289
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.
OSTI ID:
5272289
Journal Information:
Bioconjugate Chemistry; (United States), Journal Name: Bioconjugate Chemistry; (United States) Vol. 1:5; ISSN 1043-1802; ISSN BCCHE
Country of Publication:
United States
Language:
English