Identifying errors and alternate conformers and modeling multiple main-chain conformers in poorly ordered regions are overarching problems in crystallographic structure determination that have limited automation efforts and structure... more
Identifying errors and alternate conformers and modeling multiple main-chain conformers in poorly ordered regions are overarching problems in crystallographic structure determination that have limited automation efforts and structure quality. Here, we show that implementation of a full factorial designed set of standard refinement approaches, termed ExCoR (Extensive Combinatorial Refinement), significantly improves structural models compared to the traditional linear tree approach, in which individual algorithms are tested linearly and are only incorporated if the model improves. ExCoR markedly improved maps and models and reveals building errors and alternate conformations that were masked by traditional refinement approaches. Surprisingly, an individual algorithm that renders a model worse in isolation could still be necessary to produce the best overall model, suggesting that model distortion allows escape from local minima of optimization target function, here shown to be a hallmark limitation of the traditional approach. ExCoR thus provides a simple approach to improving structure determination.
Research Interests:
Research Interests:
Cofactors modulate nuclear receptor activity and impact human health and disease, yet surprisingly little is known about their transcriptional regulation. Androgen receptor trapped clone-27 (ART-27) is a cofactor that binds to androgen... more
Cofactors modulate nuclear receptor activity and impact human health and disease, yet surprisingly little is known about their transcriptional regulation. Androgen receptor trapped clone-27 (ART-27) is a cofactor that binds to androgen receptor (AR) amino terminus and modulates AR-dependent transcription. Interestingly, ART-27 displays both a cell type- and developmental stage-specific expression pattern. However, the cis-acting elements and trans-acting factors affecting ART-27 gene expression have not been elucidated. We found that ART-27 gene expression is repressed and its promoter is histone H3-K27 tri-methylated in human embryonic kidney cells, but not prostate cells, and the histone deacetylase inhibitor, trichostatin A, relieves this inhibition. The DNA response elements that control the induction of ART-27 gene expression were also characterized. The major cis-acting element corresponds to a consensus cAMP-responsive element (CRE) and binds the CRE-binding protein (CREB) as shown by EMSA and chromatin immunoprecipitation assays. Furthermore, ART-27 promoter activity is induced upon CREB overexpression. Epidermal growth factor, which activates CREB via phosphorylation, also induces ART-27 expression, whereas a reduction in CREB phosphorylation or expression blocks this induction in prostate cells. In human prostate development, both epithelial and stromal cells express CREB; however, active phosphorylated CREB is restricted to epithelial cells where ART-27 is expressed. Based on these findings, we propose a transcriptional regulatory circuit for the developmental expression of ART-27 that includes repression by chromatin modification through a trichostatin A-sensitive factor and activation upon growth factor stimulation via CREB.