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8-Bromo-adenosine diphosphoribose (br'ADP-Rib) and nicotinamide 8-bromoadenine dinucleotide (Nbr'AD') which are analogues of the coenzyme NAD' , were prepared and their liver alcohol dehydrogenase complexes studied by crystallographic methods. Nbr'AD' is active in hydrogen transport and br' ADP-Rib is a coenzyme competitive inhibitor for the enzymes liver alcohol dehydrogenase and yeast alcohol dehydrogenase. X-ray data were obtained for the complex between liver alcohol dehydrogenase and br8ADP-Rib to 0.45 nm resolution and for the liver alcohol dehydrogenase-adenosine diphosphoribose complex to 0.29-nm resolution. The conformations of these analogues were determined from the X-ray data. It was found that ADP-Rib had a conformation very similar to the corresponding part of NAD', when NAD' is bound to lactate and malate dehydrogenase.br'ADP-Rib had the same anti conformation of the adenine ring with respect to the ribose as ADP-Rib and NAD', in contrast to the syn conformation Sound in 8-bromo-adenosine. The overcrowding at the 8-position is relieved in br'ADP-Rib by having the ribose in the 2' endu conformation instead of the usual 3' endo as in ADP-Rib and NAD'.

The conformation of the polypeptide chain in horse liver alcohol dehydrogenase (EC 1.1.1.1), as well as the binding sites for some inhibitor molecules, have been determined from x-ray crystallographic data to a resolution of 2.9 A. The apoenzyme of LADH crystallizes in space-group C2221 with one subunit per asymmetric unit and cell dimensions a = 56.0 A, b = 75.2 A, and c = 181.6 A (10). The crystallographic 2-fold axis relating the two subunits of the apoenzyme molecule is not present in crystals of complexes between apoenzyme and coenzyme, which suggests that the coenzyme may induce a structural asymmetry in the chemically identical subunits. An electron density map to 5-A resolution of the apoenzyme molecule has been described (11).Here we report the structure of this molecule as deduced from an electron-density distribution at a resolution of 2.9 A. Most important is the analysis of the binding sites for some inhibitor molecules, which permits us to identify functional attributes of the enzyme structure; MATERIALS AND METHODSThe enzyme was isolated from fresh horse livers (Akeson, A. & Lundqvist, G., to be published). Preparation of crystals and heavy-atom derivatives suitable for x-ray studies has been described (10). Methods of isomorphous replacement similar to those used for other protein-structure determinations were applied to obtain the electron density map. The crystallographic data were measured at +40 on a computercontrolled Philips-Stoe four-circle diffractometer equipped with a 32 K disc storage. Data were collected to a resolution of 2.9 X from crystals of the native protein, three heavy-metal derivatives [K2Pt(CN)4, KAu(CN)2, and K2Pt(CN)4 + KAu(CN)2], and one inhibitor complex [adenosine diphosphate ribose (ADP-ribose) ]. Intensities within 4.5 A were also measured on two other inhibitor complexes: 8-Br-ADPribose and 1,10-phenanthroline. A skeletal model of the main chain was built with the Kendrew-type models, with an optical comparator (12). CONFORMATION OF THE SUBUNITIn this section we describe briefly the conformation of the polypeptide chain and some details of the subunit interaction and binding of inhibitor molecules as deduced from our electron-density maps. In the next section we will discuss some implications of this structure.The two highest features in our 2.9-A electron-density map were roughly spherical in shape and were interpreted as the two zinc atoms of the subunit. From our 5-A work we already knew the position of one of these zinc atoms and the subunit