Klebsiella pneumoniae cystathionine β-lyase – structure-functional analysis of a promising component of the enzyme-prodrug binary system

• Klebsiella pneumoniae CBL was found to catalyze thiosulfinates formation. • Recombinant CBL transforms a broad range of S-substituted L-cysteine sulfoxides. • Enzymatic conversion of the substrates into the thiosulfinates reaches up to 100 %. • Spatial structures of the holoenzyme and its complex with methiin was obtained. • The complex with methiin captured the step of α-aminoacrylate formation. In situ enzymatic production of bioactive compounds from their non-toxic precursors is a relatively new strategy in drug design. If used correctly, the enzyme-prodrug binary system allows to obtain the necessary therapeutic concentrations of drugs locally and under controlled release. In nature alliinase uses S -substituted L-cysteine sulfoxide as the key part of the defense mechanisms in plants of the Allium genus, and the resulting biologically active thiosulfinates have various health benefits. In our study we describe the recombinant enzyme cystathionine β-lyase from Klebsiella pneumoniae (kCBL) and its capability to catalyze the similar process, transforming a wide range of S -alk(en)yl- and arylalkyl-derivatives of L-cysteine sulfoxide into the corresponding thiosulfinates. The reaction proceeds with high catalytic efficiency and reaches up to 100 % conversion of the substrate. Besides a comprehensive biochemical analysis, the 3D structure of the holoenzyme and its complex with S -methyl-L-cysteine sulfoxide have been obtained at 1.9 and 2.0 Å resolution, accordingly. The binding pocket of the enzyme provides suitable environment to catalyze β-elimination reaction of the substituted L-cysteine sulfoxides of different size. To our knowledge, kCBL is the only described bacterial enzyme with such a wide substrate specificity, which makes it promising to be used as a component of the enzyme-prodrug binary system.