2-acetyl-1-pyrroline and 4-aminobutyraldehyde

Topics: Aldehydes; Betaine-Aldehyde Dehydrogenase; Odorants; Pandanaceae; Pyrroles; Real-Time Polymerase Chain Reaction

Fragrance rice (Oryza sativa) contains two isoforms of BADH, named OsBADH1 and OsBADH2. OsBADH1 is implicated in acetaldehyde oxidation in rice plant peroxisomes, while the non-functional OsBADH2 is believed to be involved in the accumulation of 2-acetyl-1-pyrroline, the major compound of aroma in fragrance rice. In the present study, site-directed mutagenesis, molecular docking and molecular dynamics simulation studies were used to investigate the substrate specificity towards Bet-ald and GAB-ald. Consistent with our previous study, kinetics data indicated that the enzymes catalyze the oxidation of GAB-ald more efficiently than Bet-ald and the OsBADH1 W172F and OsBADH2 W170F mutants displayed a higher catalytic efficiency towards GAB-ald. Molecular docking analysis and molecular dynamics simulations for the first time provided models for aldehyde substrate-bound complexes of OsBADHs. The amino acid residues, E262, L263, C296 and W461 of OsBADH1 and E260, L261, C294 and W459 of OsBADH2 located within 5 Å of the OsBADH active site mainly interacted with GAB-ald forming strong hydrogen bonds in both OsBADH isoforms. Residues W163, N164, Q294, C296 and F397 of OsBADH1-Bet-ald and Y163, M167, W170, E260, S295 and C453 of OsBADH2-Bet-ald formed the main interaction sites while E260 showed an interaction energy of -14.21 kcal/mol. Unconserved A290 in OsBADH1 and W288 in OsBADH2 appeared to be important for substrate recognition similar to that observed in PsAMADHs. Overall, the results here help to explain how two homologous rice BADHs recognize the aldehyde substrate differently, a key property to their biological role.

Topics: Acetaldehyde; Aldehydes; Betaine-Aldehyde Dehydrogenase; Catalytic Domain; Molecular Dynamics Simulation; Mutagenesis, Site-Directed; Oryza; Protein Conformation; Protein Isoforms; Pyrroles; Substrate Specificity

Betaine aldehyde dehydrogenase 2 (BADH2) is believed to be involved in the accumulation of 2-acetyl-1-pyrroline (2AP), one of the major aromatic compounds in fragrant rice. The enzyme can oxidize ω-aminoaldehydes to the corresponding ω-amino acids. This study was carried out to investigate the function of wild-type BADHs and four BADH2 mutants: BADH2_Y420, containing a Y420 insertion similar to BADH2.8 in Myanmar fragrance rice, BADH2_C294A, BADH2_E260A and BADH2_N162A, consisting of a single catalytic-residue mutation. Our results showed that the BADH2_Y420 mutant exhibited less catalytic efficiency towards γ-aminobutyraldehyde but greater efficiency towards betaine aldehyde than wild-type. We hypothesized that this point mutation may account for the accumulation of γ-aminobutyraldehyde/Δ(1)-pyrroline prior to conversion to 2AP, generating fragrance in Myanmar rice. In addition, the three catalytic-residue mutants confirmed that residues C294, E260 and N162 were involved in the catalytic activity of BADH2 similar to those of other BADHs.

Topics: Aldehydes; Betaine-Aldehyde Dehydrogenase; Biocatalysis; Kinetics; Models, Molecular; Molecular Sequence Data; Oryza; Plant Proteins; Point Mutation; Pyrroles