The Epstein-Barr Virus Oncoprotein, LMP1, Regulates the Function of SENP2, a SUMO-protease.

Epstein-Barr virus (EBV) latent membrane protein-1 (LMP1) activates numerous signal transduction pathways using its C-terminal activating regions. We reported that LMP1 increased global levels of sumoylated proteins, which aided the oncogenic nature of LMP1. Because increased protein sumoylation is detected in numerous cancers, we wanted to elucidate additional mechanisms by which LMP1 modulates the sumoylation machinery.

Crystallization, optimization and preliminary X-ray characterization of a metal-dependent PI-PLC from Streptomyces antibioticus.

A recombinant metal-dependent phosphatidylinositol-specific phospholipase C (PI-PLC) from Streptomyces antibioticus has been crystallized by the hanging-drop method with and without heavy metals. The native crystals belonged to the orthorhombic space group P222, with unit-cell parameters a=41.26, b=51.

Characterization of a randomized FRET library for protease specificity determination.

Protease specificity determination is an important first step when characterizing novel proteases. Given the large number of proteases that are known to exist from genomic sequencing efforts, we reason that sensitive, reliable, and high-throughput methods to determine protease specificity must be developed. This study describes the construction and initial characterization of a protein based FRET library using the fluorescent proteins GFP and DsRed for such a purpose.

Computational active site analysis of molecular pathways to improve functional classification of enzymes.

This study describes a method to computationally assess the function of homologous enzymes through small molecule binding interaction energy. Three experimentally determined X-ray structures and four enzyme models from ornithine cyclo-deaminase, alanine dehydrogenase, and mu-crystallin were used in combination with nine small molecules to derive a function score (FS) for each enzyme-model combination. While energy values varied for a single molecule-enzyme combination due to differences in the active sites, we observe that the binding energies for the entire pathway were proportional for each set of small molecules investigated.

X-ray structure of the R69D phosphatidylinositol-specific phospholipase C enzyme: insight into the role of calcium and surrounding amino acids in active site geometry and catalysis.

Phosphatidylinositol-specific phospholipase Cs (PLCs) are a family of phosphodiesterases that catalyze the cleavage of the P-O bond via transesterification using the internal hydroxyl group of the substrate as a nucleophile, generating the five-membered cyclic inositol phosphate as an intermediate or product. To better understand the role of calcium in the catalytic mechanism of PLCs, we have determined the X-ray crystal structure of an engineered PLC enzyme from Bacillus thuringiensis to 2.1 A resolution.

Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline.

Structural genomics is emerging as a principal approach to define protein structure-function relationships. To apply this approach on a genomic scale, novel methods and technologies must be developed to determine large numbers of structures. We describe the design and implementation of a high-throughput structural genomics pipeline and its application to the proteome of the thermophilic bacterium Thermotoga maritima.