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Western University

Biochemistry Publications

Protein Binding

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Full-Text Articles in Life Sciences

Discovery And Validation Of Information Theory-Based Transcription Factor And Cofactor Binding Site Motifs., Ruipeng Lu, Eliseos J Mucaki, Peter K Rogan Mar 2017

Discovery And Validation Of Information Theory-Based Transcription Factor And Cofactor Binding Site Motifs., Ruipeng Lu, Eliseos J Mucaki, Peter K Rogan

Biochemistry Publications

Data from ChIP-seq experiments can derive the genome-wide binding specificities of transcription factors (TFs) and other regulatory proteins. We analyzed 765 ENCODE ChIP-seq peak datasets of 207 human TFs with a novel motif discovery pipeline based on recursive, thresholded entropy minimization. This approach, while obviating the need to compensate for skewed nucleotide composition, distinguishes true binding motifs from noise, quantifies the strengths of individual binding sites based on computed affinity and detects adjacent cofactor binding sites that coordinate with the targets of primary, immunoprecipitated TFs. We obtained contiguous and bipartite information theory-based position weight matrices (iPWMs) for 93 sequence-specific TFs, …


Tethering Polypeptides Through Bifunctional Peg Cross-Linking Agents To Probe Protein Function: Application To Atp Synthase., Daniel J Cipriano, Stanley D Dunn Nov 2008

Tethering Polypeptides Through Bifunctional Peg Cross-Linking Agents To Probe Protein Function: Application To Atp Synthase., Daniel J Cipriano, Stanley D Dunn

Biochemistry Publications

Chemical crosslinking mediated by short bifunctional reagents has been widely used for determining physical relationships among polypeptides in multisubunit proteins, but less often for functional studies. Here we introduce the approach of tethering polypeptides by using bifunctional reagents containing a lengthy, flexible PEG linker as a form of crosslinking especially suited to functional analyses. The rotary molecular motor ATP synthase was used as a model subject. Single cysteine residues were introduced into selected positions of ATP synthase epsilon subunit, a component of the rotor subcomplex of the enzyme, and the unrelated maltose binding protein (MBP), then the two purified recombinant …


Crystal Structures Of The Streptomyces Coelicolor Tetr-Like Protein Actr Alone And In Complex With Actinorhodin Or The Actinorhodin Biosynthetic Precursor (S)-Dnpa., A R Willems, K Tahlan, T Taguchi, K Zhang, Z Z Lee, K Ichinose, M S Junop, J R Nodwell Mar 2008

Crystal Structures Of The Streptomyces Coelicolor Tetr-Like Protein Actr Alone And In Complex With Actinorhodin Or The Actinorhodin Biosynthetic Precursor (S)-Dnpa., A R Willems, K Tahlan, T Taguchi, K Zhang, Z Z Lee, K Ichinose, M S Junop, J R Nodwell

Biochemistry Publications

Actinorhodin, an antibiotic produced by Streptomyces coelicolor, is exported from the cell by the ActA efflux pump. actA is divergently transcribed from actR, which encodes a TetR-like transcriptional repressor. We showed previously that ActR represses transcription by binding to an operator from the actA/actR intergenic region. Importantly, actinorhodin itself or various actinorhodin biosynthetic intermediates can cause ActR to dissociate from its operator, leading to derepression. This suggests that ActR may mediate timely self-resistance to an endogenously produced antibiotic by responding to one of its biosynthetic precursors. Here, we report the structural basis for this precursor-mediated derepression with crystal structures of …


The Stator Complex Of The A1a0-Atp Synthase--Structural Characterization Of The E And H Subunits., Erik Kish-Trier, Lee-Ann K Briere, Stanley D Dunn, Stephan Wilkens Jan 2008

The Stator Complex Of The A1a0-Atp Synthase--Structural Characterization Of The E And H Subunits., Erik Kish-Trier, Lee-Ann K Briere, Stanley D Dunn, Stephan Wilkens

Biochemistry Publications

Archaeal ATP synthase (A-ATPase) is the functional homolog to the ATP synthase found in bacteria, mitochondria and chloroplasts, but the enzyme is structurally more related to the proton-pumping vacuolar ATPase found in the endomembrane system of eukaryotes. We have cloned, overexpressed and characterized the stator-forming subunits E and H of the A-ATPase from the thermoacidophilic Archaeon, Thermoplasma acidophilum. Size exclusion chromatography, CD, matrix-assisted laser desorption ionization time-of-flight mass spectrometry and NMR spectroscopic experiments indicate that both polypeptides have a tendency to form dimers and higher oligomers in solution. However, when expressed together or reconstituted, the two individual polypeptides interact with …