Cloning of cDNAs for human aldehyde dehydrogenases 1 and 2.

Abstract

Partial cDNA clones encoding human cytosolic aldehyde dehydrogenase (ALDH1) and mitochondrial aldehyde dehydrogenase (ALDH2) were isolated from a human liver cDNA library constructed in phage Agtll. The expression library was screened by using rabbit antibodies against ALDH1 and ALDH2. Positive clones thus obtained were subsequently screened with mixed synthetic oligonudeotides compatible with peptide sequences of ALDH1 and ALDH2. One of the positive clones for ALDH1 contained an insertion of 1.6 kilobase pairs (kbp). The insert encoded 340 amino acid residues and had a 3' noncoding region of 538 bp and a poly(A) segment. The amino acid sequence deduced from the cDNA sequence coincided with the reported amino acid sequence of human ALDH1 [Hempel, J., von Bahr-Lindstrom, H. & Jornvall, H. (1984) Eur. J. Biochem. 141, 21-35], except that valine at position 161 in the previous amino acid sequence study was found to be isoleucine in the deduced sequence. Since the amino acid sequence of ALDH2 was unknown, 33 tryptic peptides of human ALDH2 were isolated and sequenced. Based on the amino acid sequence data thus obtained, a mixed oligonucleotide probe was prepared. Two positive clones, AALDH2-21 and AALDH2-36, contained the same insert of 1.2 kbp. Another done, AALDH2-22, contained an insert of 1.3 kbp. These two inserts contained an overlap region of 0.9 kbp. The combined cDNA contained a sequence that encodes 399 amino acid residues, a chain-termination codon, a 3' untranslated region of 403 bp, and a poly(A) segment. The deduced amino acid sequence was compatible with the amino acid sequences of the tryptic peptides. The degree of homology between human ALDH1 and ALDH2 is 66% for the coding regions of their cDNAs and 69% at the protein level. No significant homology was found in their 3' untranslated regions.Liver aldehyde dehydrogenase (ALDH; aldehyde:NAD+ oxidoreductase, EC 1.2.1.3) is considered to play a major role in alcohol metabolism. Two major and several minor isozymes exist in the livers of mammals, including man.

Xenopus Cytosolic Thyroid Hormone-binding Protein (xCTBP) Is Aldehyde Dehydrogenase Catalyzing the Formation of Retinoic Acid

“…The information about the sequence involved in the [ 125 I]T 3 -binding site was obtained by the identification of the peptide with radioactivity after digestion of the affinity labeled xCTBP with lysyl endopeptidase. Protein sequencing revealed that the peptide (K)LADLVERDRLILSTM-, which corresponded well to 91-106 residues of human ALDH1 enzyme (25) and to 92-107 residues of xALDH1-I and -II enzymes Table I. xCTBP catalyzed the conversion of acetaldehyde into acetic acid or retinal into RA at a linear rate for at least 1-2 min at 1.85 g of the purified protein.…”

“…Screening of cDNA Library and Sequence Analysis-An hepatic cDNA library was screened with 32 P-radiolabeled human ALDH1 cDNA (25). The entire sequences of the two cDNAs were determined for both strands by the method of Sanger et al (28).…”

“…Toward this aim, we have cloned two cDNAs encoding ALDH1 from a Xenopus hepatic cDNA library with human ALDH1 cDNA as the probe (25), determined their nucleotide sequences, and examined both the enzyme and T 3 binding activities using the recombinant proteins expressed in Escherichia coli. Deduced amino acid sequences and studies of their activities clearly showed that the two translated products are ALDH1 with T 3 binding activity.…”

Polymorphisms of Alcohol Metabolizing Enzymes in Indigenous Mexican Population: Unusual High Frequency of CYP2E1*c2 Allele

“…ALDH2*2 mutant allele is characterized by the loss of a Ksp632I restriction site and a resulting enzyme with little or no catalytic activity (Crabb et al, 1989;Hsu et al, 1985).…”

Alcohol and Treatment of Diabetes

“…Hsu et al, on the other hand, established the association of the oriental flush reaction with genetically determined absence of a mitochondria1 isoenzyme ALDH2, which is responsible for most acetaldehyde metabolism (33). There is no absence of this enzyme in Caucasians who exhibit an alcohol flush.…”