Systems-based design of bi-ligand inhibitors of oxidoreductases: filling the chemical proteomic toolbox.

Genomics-driven growth in the number of enzymes of unknown function has created a need for better strategies to characterize them. Since enzyme inhibitors have traditionally served this purpose, we present here an efficient systems-based inhibitor design strategy, enabled by bioinformatic and NMR structural developments. First, we parse the oxidoreductase gene family into structural subfamilies termed pharmacofamilies, which share pharmacophore features in their cofactor binding sites.

UBA domains mediate protein-protein interactions between two DNA damage-inducible proteins.

The Saccharomyces cerevisiae genes RAD23 and DDI1 were identified in a screen for multicopy suppressors of the temperature-sensitivity of a mutant allele of S. cerevisiae PDS1. Pds1 is a regulator of anaphase that needs to accumulate and then be degraded by the ubiquitin-proteasome pathway at the metaphase-anaphase transition for cells to progress normally through mitosis.

UBA domains of DNA damage-inducible proteins interact with ubiquitin.

Rad23 is a highly conserved protein involved in nucleotide excision repair (NER) that associates with the proteasome via its N-terminus. Its C-terminal ubiquitin-associated (UBA) domain is evolutionarily conserved from yeast to humans. However, the cellular function of UBA domains is not completely understood.

Dosage suppressors of pds1 implicate ubiquitin-associated domains in checkpoint control.

In budding yeast, anaphase initiation is controlled by ubiquitin-dependent degradation of Pds1p. Analysis of pds1 mutants implicated Pds1p in the DNA damage, spindle assembly, and S-phase checkpoints. Though some components of these pathways are known, others remain to be identified.

Microinjection of anti-p21 antibodies induces senescent Hs68 human fibroblasts to synthesize DNA but not to divide.

Replicative senescence is characterized by irreversible growth arrest and has been defined by four genetic complementation groups. One of these groups is associated with the predominance of underphosphorylated, growth-suppressive retinoblastoma tumor suppressor protein (pRb). Although certain members of the cyclin-dependent kinase (cdk)/cyclin family, some of which phosphorylate pRb, are underexpressed in senescent cells, others are expressed but inactive.