Photo-reversible binding of thrombin to avidin by means of a photolabile inhibitor.
Human alpha-thrombin and Factor Xa 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 light. On irradiation with light of wavelength 366 nm for 6 min, however, up to 80% of pre-inhibition activity was regained. This photo-deacylation of the modified enzyme results in the formation of active enzyme and a coumarin by-product (3). In addition, the acyl enzyme that results from incubation of 1c with thrombin was capable of binding to avidin, both immobilized and free in solution. Furthermore, the complex formed between the thrombin acyl enzyme (2c) and avidin was capable of binding to immobilized biotin.
Porter
N A
NA
Department of Chemistry, Duke University, Durham, NC 27708, USA.
Bush
K A
KA
Kinter
K S
KS
D000215:Acylation; D001360:Avidin / Q000378:metabolism*; D001665:Binding Sites; D001710:Biotin / Q000737:chemistry*; D002934:Cinnamates / Q000737:chemistry*; D015951:Factor Xa / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008956:Models, Chemical; D010777:Photochemistry; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*
D002934:Cinnamates; D001360:Avidin; D001710:Biotin; D013917:Thrombin; D015951:Factor Xa