Cloning, expression, and characterization of an aldehyde dehydrogenase from Escherichia coli K-12 that utilizes 3-Hydroxypropionaldehyde as a substrate.
3-Hydroxypropionaldehyde (3-HPA), an intermediary compound of glycerol metabolism in bacteria, serves as a precursor to 3-Hydroxypropionic acid (3-HP), a commercially valuable platform chemical. To achieve the effective conversion of 3-HPA to 3-HP, an aldH gene encoding an aldehyde dehydrogenase in Escherichia coli K-12 (AldH) was cloned, expressed, and characterized for its properties. The recombinant AldH exhibited broad substrate specificity for various aliphatic and aromatic aldehydes. AldH preferred NAD+ over NADP+ as a cofactor for the oxidation of most aliphatic aldehydes tested. Among the aldehydes used, the specific activity was highest (38.1 U mg(-1) protein) for 3-HPA at pH 8.0 and 37 degrees C. The catalytic efficiency (kcat) and the specificity constant (kcat/Km) for 3-HPA in the presence of NAD+ were 28.5 s(-1) and 58.6x10(3) M(-1) s(-1), respectively. The AldH activity was enhanced in the presence of disulfide reductants such as dithiothreitol (DTT) or 2-mercaptoethanol, while several metal ions, particularly Hg2+, Ag+, Cu2+, and Zn2+, inhibited AldH activity. This study illustrates that AldH is a potentially useful enzyme in converting 3-HPA to 3-HP.
Jo
Ji-Eun
JE
Department of Chemical and Biochemical Engineering, Pusan National University, Busan, 609-735, Republic of Korea.
Mohan Raj
Subramanian
S
Rathnasingh
Chelladurai
C
Selvakumar
Edwardraja
E
Jung
Woo-Chel
WC
Park
Sunghoon
S
D000444:Aldehyde Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D004795:Enzyme Stability; D048168:Escherichia coli K12 / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015870:Gene Expression; D005985:Glyceraldehyde / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D019344:Lactic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D008969:Molecular Sequence Data; D011407:Propane / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D003067:Coenzymes; D029968:Escherichia coli Proteins; C047158:3-hydroxypropionaldehyde; D019344:Lactic Acid; D005985:Glyceraldehyde; C031601:hydracrylic acid; D000444:Aldehyde Dehydrogenase; D011407:Propane