Whole-genome characterization of human and simian immunodeficiency virus intrahost diversity by ultradeep pyrosequencing.

Rapid evolution and high intrahost sequence diversity are hallmarks of human and simian immunodeficiency virus (HIV/SIV) infection. Minor viral variants have important implications for drug resistance, receptor tropism, and immune evasion. Here, we used ultradeep pyrosequencing to sequence complete HIV/SIV genomes, detecting variants present at a frequency as low as 1%.

Homogenous fluorescent assays for characterizing small-molecule activators of AMP-activated protein kinase (AMPK).

AMP activated protein kinase (AMPK) is a key regulator of cellular metabolism. AMPK activity is modulated in part by binding of AMP to the gamma-subunit of the kinase, which increases the activity of the catalytic alpha-subunit. Because increased AMPK activity in the liver and in skeletal muscle leads to increased fatty acid oxidation and decreased cholesterol and fatty acid biosynthesis, activators of AMPK are being sought for treatment of type-2 diabetes and other metabolic disorders.

Definition of the extended substrate specificity determinants for beta-tryptases I and II.

Tryptases betaI and betaII were heterologously expressed and purified in yeast to functionally characterize the substrate specificity of each enzyme. Three positional scanning combinatorial tetrapeptide substrate libraries were used to determine the primary and extended substrate specificity of the proteases. Both enzymes have a strict primary preference for cleavage after the basic amino acids, lysine and arginine, with only a slight preference for lysine over arginine.

Recombinant human mast cell tryptase beta: stable expression in Pichia pastoris and purification of fully active enzyme.

Human mast cell tryptase beta (EC 3.4.21.

Factors that modify the molecular size of phospholamban, the 23,000-dalton cardiac sarcoplasmic reticulum phosphoprotein.

Phospholamban, a 23,000-dalton phosphoprotein present in cardiac muscle sarcoplasmic reticulum vesicles is quantitatively dissociated into three smaller sized phosphorylated components of 11,000, 15,000, and 20,000 daltons when sarcoplasmic reticulum, solubilized in 1% (w/v) sodium dodecyl sulfate and 1 mM MgCl2, is heated at 71 degrees C or greater for 2 min. This dissociation is inhibited by Mg2+ (50% at approximately 5 mM). The 23,000-dalton phosphoprotein reformed from the 11,000-, 15,000-, and 20,000-dalton phosphorylated components when phosphorylated sarcoplasmic reticulum that had been boiled in 1% sodium dodecyl sulfate and 1 MM MgCl2 was stored for 1 week at -70 degrees C.