Three-dimensional structures of the three human class I alcohol dehydrogenases

Abstract

In contrast with other animal species, humans possess three distinct genes for class I alcohol dehydrogenase and show polymorphic variation in the ADH1B and ADH1C genes. The three class I alcohol dehydrogenase isoenzymes share approximately 93% sequence identity but differ in their substrate specificity and their developmental expression. We report here the first three-dimensional structures for the ADH1A and ADH1C*2 gene products at 2.5 and 2.0 A, respectively, and the structure of the ADH1B*1 gene product in a binary complex with cofactor at 2.2 A. Not surprisingly, the overall structure of each isoenzyme is highly similar to the others. However, the substitution of Gly for Arg at position 47 in the ADH1A isoenzyme promotes a greater extent of domain closure in the ADH1A isoenzyme, whereas substitution at position 271 may account for the lower turnover rate for the ADH1C*2 isoenzyme relative to its polymorphic variant, ADH1C*1. The substrate-binding pockets of each isoenzyme possess a unique topology that dictates each isoenzyme's distinct but overlapping substrate preferences. ADH1*B1 has the most restrictive substrate-binding site near the catalytic zinc atom, whereas both ADH1A and ADH1C*2 possess amino acid substitutions that correlate with their better efficiency for the oxidation of secondary alcohols. These structures describe the nature of their individual substrate-binding pockets and will improve our understanding of how the metabolism of beverage ethanol affects the normal metabolic processes performed by these isoenzymes.

Fig. 1.

Ribbon diagrams of the aligned human class I dimeric isoenzymes. (A) The ADH1C*2 isoenzyme (blue) was aligned with the ADH1B*1 isoenzyme (green) by using the α-carbon atoms in their respective coenzyme-binding domains (residues 176–322 in both subunits). (B) The ADH1A isoenzyme (red) was aligned with the ADH1B*1 isoenzyme (green) by using the same procedure as for the ADH1C*2 isoenzyme. The ribbon diagrams were produced using the programs MOLSCRIPT and Raster3D (Bacon and Anderson 1988; Kraulis 1991; Merrit and Murphy 1994).

Fig. 2.

Stereodiagrams of the substrate-binding sites of the human class I isoenzymes. The identical alignment procedure used in Fig. 1 ▶ was used here to minimize differences in the cofactor positioning between the aligned substrate-binding sites. (A) The substrate-binding site in ADH1B*1 (green) overlayed on the corresponding region of the ADH1C*2 substrate-binding site. (Zn) Active site zinc atom; (Wat) zinc-bound water. Individual residues are labeled with their sequence identifiers. (B) The substrate-binding site of ADH1A (red) overlayed on the substrate-binding site of ADH1B*1 (green). The active site zinc is labeled identically as in Fig. 2A ▶, and the position of the partially occupied 4-iodopyrazole molecule is shown in blue and labeled 4IP. The figure was generated using the program MOLSCRIPT (Kraulis 1991).

Fig. 2.

Stereodiagrams of the substrate-binding sites of the human class I isoenzymes. The identical alignment procedure used in Fig. 1 ▶ was used here to minimize differences in the cofactor positioning between the aligned substrate-binding sites. (A) The substrate-binding site in ADH1B*1 (green) overlayed on the corresponding region of the ADH1C*2 substrate-binding site. (Zn) Active site zinc atom; (Wat) zinc-bound water. Individual residues are labeled with their sequence identifiers. (B) The substrate-binding site of ADH1A (red) overlayed on the substrate-binding site of ADH1B*1 (green). The active site zinc is labeled identically as in Fig. 2A ▶, and the position of the partially occupied 4-iodopyrazole molecule is shown in blue and labeled 4IP. The figure was generated using the program MOLSCRIPT (Kraulis 1991).

Fig. 3.

Stereodiagrams of the coenzyme-binding site residues in ADH1A and ADH1C*2 near the positions or residues 47 and 271, respectively. (A) The structure of the ADH1A isoenzyme (red) in the vicinity of Gly 47 overlayed onto the same region of the ADH1B*1 isoenzyme (green). (Wat1, Wat2, Wat3) Positions of the three water molecules recruited to take the place of Arg 47. Three additional ordered water molecules (not labeled) that are common between all three human ADH1 isoenzymes also are shown. The alignment procedure was identical to the one used in Fig. 1 ▶. (B) The structure of the ADH1C*2 isoenzyme (blue) in the vicinity of Gln 271 overlayed onto the similar region of the ADH1B*1 isoenzyme. Three water molecules common to these two structures are shown near Lys 228 and the N6′ atom of the adenine ring. In addition, an ordered water molecule unique to this region of the ADH1C*2 also is shown positioned between Asp 273 and Gln 271. The figures were generated using MOLSCRIPT (Kraulis 1991).

Fig. 3.

Stereodiagrams of the coenzyme-binding site residues in ADH1A and ADH1C*2 near the positions or residues 47 and 271, respectively. (A) The structure of the ADH1A isoenzyme (red) in the vicinity of Gly 47 overlayed onto the same region of the ADH1B*1 isoenzyme (green). (Wat1, Wat2, Wat3) Positions of the three water molecules recruited to take the place of Arg 47. Three additional ordered water molecules (not labeled) that are common between all three human ADH1 isoenzymes also are shown. The alignment procedure was identical to the one used in Fig. 1 ▶. (B) The structure of the ADH1C*2 isoenzyme (blue) in the vicinity of Gln 271 overlayed onto the similar region of the ADH1B*1 isoenzyme. Three water molecules common to these two structures are shown near Lys 228 and the N6′ atom of the adenine ring. In addition, an ordered water molecule unique to this region of the ADH1C*2 also is shown positioned between Asp 273 and Gln 271. The figures were generated using MOLSCRIPT (Kraulis 1991).