(1) CCRLC Daresbury Laboratory, Warrington WA4 4AD, UK
(2) EMBL c/o ILL, F-38043 Grenoble, CEDEX, France.
(3) Institute of Biochemistry RAS, 34 Leninsky pr, 117071 Moscow, Russia
Fumarase and delta-crystallin (DCRY) belong to a superfamily of metabolic enzymes which are all active as homotetramers with approximate molecular weight of 200 kDa. Members of the superfamily also include aspartase, adenylosuccinase, argininosuccinase and 3-carboxy-cis,cis-muconate lactonising enzyme (CMLE). These enzymes catalize a similar trans-elimination reaction with the fumarate as a final product [1]. The eye lens protein DCRY has no catalitic activity. However, it shares approximately 90% sequence identity with argininosuccinase, indicating that it is an example of a "hijacked" enzyme. The avian lens DCRY was the first structure from the family to be determined by the X-ray methods [2]. The X-ray structure of yeast fumarase [3] has recently been solved by the molecular replacement method using the 2.4 Å synchrotron data collected at 100 K and the coordinates of the X-ray structure of the E.coli enzyme as a model [4]. The monomer of fumarase, containing around 480 amino acids, consists of three highly alpha-helical domains. The three domains form an elongated structure which resembles a dumb-bell. The four monomers are arranged in a tetramer with point group symmetry 222. The structure comparison with DCRY reveals very high similarity between the central large domains of the monomers. The packing of four subunits in the tetramer is also very similiar. The active site in fumarase is built up using the residues from three subunits. Most of these residues are highly conserved within the superfamily. Examining the fumarase active site cleft shows that the residues responsible for the catalytic activity occupy homologous positions in the putative active site of the DCRY structure. At this stage it is difficult to explain why DCRY has lost its enzymatic activity. Further details of the comparison of the active sites will be presented.
Acknowledgements
We thank C.Slingsby and A.Simpson ( Birkbeck Colledge, London Univ.) for the coordinates of delta-crystallin and L.Banaszak and T.Weaver (Univ. of Minneso- ta) for the coordinates of fumarase E.coli.
References
[1] Woods, S.A. et al , FEMS Microbiol.Lett. (1988) 51:181-186
[2] Simpson, A. et al, Nature, Struct.Biol. (1994) 1:724-733
[3] Keruchenko, J.S. et al, Biochim.Biophys.Acta (1992) 1122,85-92
[4] Weaver, T.M. et al, Nature, Struct.Biol. (1995) 2:654-662