PHOTO-REVERSIBLE BINDING OF THROMBIN TO AVIDIN BY MEANS OF A PHOTOLABILE INHIBITOR

Human alpha-thrombin and Factor X, were acylated at their active site serine hydroxyls with biotin-substituted cinnamate derivatives 1b-1c. These acyl enzymes (2) showed no enzyme activity in the absence of lig ht. On irradiation with light of wavelength 366 nm for 6 min, however, up to 80% of pre-inhibition activity was regained. This photo-deacyla tion of the modified enzyme results in the formation of active enzyme and a coumarin by-product (3). In addition, the acyl enzyme that resul ts from incubation of Ic with thrombin was capable of binding to avidi n, both immobilized and free in solution. Furthermore, the complex for med between the thrombin acyl enzyme (2c) and avidin was capable of bi nding to immobilized biotin. (C) 1997 Elsevier Science S.A.