Crystal structures of human gephyrin and plant Cnx1 G domains: Comparativeanalysis and functional implications

The molybdenum cofactor (Moco) consists of a unique and conserved pterin de rivative, usually referred to as molybdopterin (MPT), which coordinates the essential transition metal molybdenum (Mo). Moco is required for the enzym atic activities of all Mo-enzymes, with the exception of nitrogenase and is synthesized by an evolutionary old multi-step pathway that is dependent on the activities of at least six gene products. In eukaryotes, the final ste p of Moco biosynthesis, i.e. transfer and insertion of Mo into MPT, is cata lyzed by the two-domain proteins Cnx1 in plants and gephyrin in mammals. Ge phyrin is ubiquitously expressed, and was initially found in the central ne rvous system, where it is essential for clustering of inhibitory neurorecep tors in the postsynaptic membrane. Gephyrin and Cnx1 contain at least two f unctional domains (E and G) that are homologous to the Escherichia coli pro teins MoeA and MogA, the atomic structures of which have been solved recent ly. Here, we present the crystal structures of the N-terminal human gephyri n G domain (Geph-G) and the C-terminal Arabidopsis thaliana Cnx1 G domain ( Cnx1-G) at 1.7 and 2.6 Angstrom resolution, respectively, These structures are highly similar and compared to MogA reveal four major differences in th eir three-dimensional structures: (1) In Geph-G and Cnx1-G an additional a- helix is present between the first beta -strand and alpha -helix of MogA. ( 2) The loop between alpha2 and beta2 undergoes conformational changes in al l three structures. (3) A beta -hairpin loop found in MogA is absent from G eph-G and Cnx1-G. (4) The C terminus of Geph-G follows a different path fro m that in MogA. Based on the structures of the eukaryotic proteins and thei r comparisons with E. coli MogA, the predicted binding site for MPT has bee n further refined. In addition, the characterized alternative splice varian ts of gephyrin are analyzed in the context of the three-dimensional structu re of Geph-G. (C) 2001 Academic Press.