Structure of human cystathionine beta-synthase: a unique pyridoxal 5'-phosphate-dependent heme protein

Abstract

Cystathionine beta-synthase (CBS) is a unique heme- containing enzyme that catalyzes a pyridoxal 5'-phosphate (PLP)-dependent condensation of serine and homocysteine to give cystathionine. Deficiency of CBS leads to homocystinuria, an inherited disease of sulfur metabolism characterized by increased levels of the toxic metabolite homocysteine. Here we present the X-ray crystal structure of a truncated form of the enzyme. CBS shares the same fold with O-acetylserine sulfhydrylase but it contains an additional N-terminal heme binding site. This heme binding motif together with a spatially adjacent oxidoreductase active site motif could explain the regulation of its enzyme activity by redox changes.

Fig. 1. Transsulfuration pathway.

Fig. 2. Overall structure of truncated CBS. (A) Topology of the fold in CBS. Above, the C-terminal domain (with the first two strands of the β-sheet formed by the N-terminal residues); below, the N-terminal domain. Both domains are of the type α/β and contain a central β-sheet surrounded by several α-helices. Strand 1 (red) is part of the C-terminal β-sheet of the other monomer in the dimer. (B) Stereo drawing showing the overall fold of CBS with every twentieth residue labeled. (C) Schematic representation of the tertiary fold of a dimer of CBS. The central β-sheets are colored in blue and the surrounding α-helices are colored in magenta. In ball-and-stick representation and marked by a black box are the active site PLP, the heme and the oxidoreductase motif. The view is down the non-crystallographic 2-fold axis, which relates the two subunits of the dimer.

Fig. 3. Structural details of CBS. (A) The active site region of CBS (gray) in a superposition with the active site of OASS (green). The sequences as well as the structure of the two enzymes are very similar. A superposition of the 25 structurally most conserved residues yields an r.m.s.d. of 1.6 Å of their C_α positions. The substrate analog of OASS methionine indicates the probable binding mode of the first substrate serine and also the region where the second substrate homocysteine is expected to bind. (B) The heme binding site of CBS with heme in green and the surrounding residues in gray. The two residues His65 and Cys52 are the ligands to the heme iron (dark red). The difference density for the cofactor is shown in red contoured at 3.5σ. (C) The oxidoreductase motif in ball-and-stick representation and (D) in a superposition with the structure of glutaredoxin. The structure of CBS is in gray, the one of glutaredoxin in green. The overall topology is very similar, but the active site motif in CBS is switched to the other helix compared with glutaredoxin.