Structure-templated predictions of novel protein interactions from sequence information.

The multitude of functions performed in the cell are largely controlled by a set of carefully orchestrated protein interactions often facilitated by specific binding of conserved domains in the interacting proteins. Interacting domains commonly exhibit distinct binding specificity to short and conserved recognition peptides called binding profiles. Although many conserved domains are known in nature, only a few have well-characterized binding profiles. Here, we describe a novel predictive method known as domain-motif interactions from structural topology (D-MIST) for elucidating the binding profiles of interacting domains. A set of domains and their corresponding binding profiles were derived from extant protein structures and protein interaction data and then used to predict novel protein interactions in yeast. A number of the predicted interactions were verified experimentally, including new interactions of the mitotic exit network, RNA polymerases, nucleotide metabolism enzymes, and the chaperone complex. These results demonstrate that new protein interactions can be predicted exclusively from sequence information.

A novel gamma-hydroxybutyrate dehydrogenase: identification and expression of an Arabidopsis cDNA and potential role under oxygen deficiency.

In plants, gamma-aminobutyrate (GABA), a non-protein amino acid, accumulates rapidly in response to a variety of abiotic stresses such as oxygen deficiency. Under normoxia, GABA is catabolized to succinic semialdehyde and then to succinate with the latter reaction being catalyzed by succinic semialdehyde dehydrogenase (SSADH). Complementation of an SSADH-deficient yeast mutant with an Arabidopsis cDNA library enabled the identification of a novel cDNA (designated as AtGH-BDH for Arabidopsis thaliana gamma-hydroxybutyrate dehydrogenase), which encodes a 289-amino acid polypeptide containing an NADP-binding domain. Constitutive expression of AtGHBDH in the mutant yeast enabled growth on 20 mm GABA and significantly enhanced the cellular concentrations of gamma-hydroxybutyrate, the product of the GHDBH reaction. These data confirm that the cDNA encodes a polypeptide with GHBDH activity. Arabidopsis plants subjected to flooding-induced oxygen deficiency for up to 4 h possessed elevated concentrations of gamma-hydroxybutyrate as well as GABA and alanine. RNA expression analysis revealed that GHBDH transcription was not up-regulated by oxygen deficiency. These findings suggest that GHBDH activity is regulated by the supply of succinic semialdehyde or by redox balance. It is proposed that GHBDH and SSADH activities in plants are regulated in a complementary fashion and that GHBDH and gamma-hydroxybutyrate function in oxidative stress tolerance.