Crystal structure of the plant PPC decarboxylase AtHAL3a complexed with an ene-thiol reaction intermediate.
The Arabidopsis thaliana protein AtHAL3a decarboxylates 4'-phosphopantothenoylcysteine to 4'-phosphopantetheine, a step in coenzyme A biosynthesis. Surprisingly, this decarboxylation reaction is carried out as an FMN-dependent redox reaction. In the first half-reaction, the side-chain of the cysteine residue of 4'-phosphopantothenoylcysteine is oxidised and the thioaldehyde intermediate decarboxylates spontaneously to the 4'-phosphopantothenoyl-aminoethenethiol intermediate. In the second half-reaction this compound is reduced to 4'-phosphopantetheine and the FMNH(2) cofactor is re-oxidised. The active site mutant C175S is unable to perform this reductive half-reaction. Here, we present the crystal structure of the AtHAL3a mutant C175S in complex with the reaction intermediate pantothenoyl-aminoethenethiol and FMNH(2). The geometry of binding suggests that reduction of the C(alpha)=C(beta) double bond of the intermediate can be performed by direct hydride-transfer from N5 of FMNH(2) to C(beta) of the aminoethenethiol-moiety supported by a protonation of C(alpha) by Cys175. The binding mode of the substrate is very similar to that previously observed for a pentapeptide to the homologous enzyme EpiD that introduces the aminoethenethiol-moiety as final reaction product at the C terminus of peptidyl-cysteine residues. This finding further supports our view that these homologous enzymes form a protein family of homo-oligomeric flavin-containing cysteine decarboxylases, which we have termed HFCD family.
Steinbacher
Stefan
S
Max-Planck-Institut für Biochemie, Abteilung für Strukturforschung, Am Klopferspitz 18a, D-82152 Martinsried, Germany. steinbac@biochem.mpg.de
Hernández-Acosta
Pilar
P
Bieseler
Bastian
B
Blaesse
Michael
M
Huber
Robert
R
Culiáñez-Macià
Francisco Antonio
FA
Kupke
Thomas
T
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002262:Carboxy-Lyases / Q000737:chemistry; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D055672:Static Electricity; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds / Q000378:metabolism*
D029681:Arabidopsis Proteins; C404262:AtHAL3 protein, Arabidopsis; D013438:Sulfhydryl Compounds; D010088:Oxidoreductases; C075971:epidermin decarboxylase; D002262:Carboxy-Lyases