Studies with mechanism-based inactivators of lysine .epsilon.-transaminase from Achromobacter liquidum

Abstract

Analogs of lysine containing a 4,5-acetylenic linkage (lysyne) or a cis- or trans-4,5-olefinic linkage (lysenes) function as substrates for a homogeneous L-lysine .epsilon.-transaminase from A. liquidum but partition between transamination and time-dependent inactivation. The partition ratio is lowest for lysyne (40 per inactivation event) and higher for trans-lysene (160 per inactivation event) and the cis-lysene transaminates 1600 times per inactivation event. cis-Lysene yields .alpha.-picolinate as a detectable accumulating product, presumably from cyclization of initial 6-aldehyde to dihydropicolinate and spontaneous autooxidation. The trans isomer also yields some picolinate as an identifiable product. The product from the few lysyne turnovers is as yet unknown but has strong absorbance at 318 nm. The inactive enzyme species from all 3 lysine analogs slowly (overnight) regain full activity after gel filtration chromatography and dialysis, suggesting reversal of the initial adduct-forming reaction. Initial studies with partially purified pseudomonad lysine .alpha.-racemase show .alpha.-3H incorporation from 3H2O but no inactivation, consistent with the expectation that these lysine analogs could act readily as mechanism-based inactivators for pyridoxal P enzymes which act at the .epsilon.- but not the .alpha.-C of lysine.