The melting behavior of a DNA fragment carrying the mouse beta maj-globin promoter was investigated as a means of establishing procedures for separating DNA fragments differing by any single base substitution using the denaturing gradient gel electrophoresis procedure of Fischer and Lerman (1,2). We find that attachment of a 300 base pair GC-rich DNA sequence, termed a GC-clamp, to a 135 bp DNA...

Sat-RNA is one of several replicating satellite RNAs which have been isolated from RNA encapsidated in cucumber mosaic virus (CMV) and which are totally dependent on CMV for replication. The 336 residue sequence of Sat-RNA obtained using the dideoxynucleotide chain termination and partial enzymic digestion procedures shows only a few short stretches (up to 11 residues) of sequence homology with...

Using both quasi-elastic and fixed-window neutron spectroscopy, we study the dynamics of highly concentrated aqueous protein solutions of bovine serum albumin around the denaturing transition. For the temperature range 280 K < T < 370 K, the total mean-squared displacement hu2i is recorded. Below and above the denaturing, we observe that hu2i increases monotonically with T, but at the denaturin...

We have developed an electrophoretic method to determine the size of a protein—DNA complex using the same binding and electrophoresis conditions as in an electrophoretic mobility shift assay (EMSA). The method is an adaptation of that of Ferguson (1), originally developed to determine the size of native proteins in non-denaturing gel systems. In SDS polyacrylamide gel electrophoresis, associati...

The aim of this study was to optimize methods for quantifying 13 uridine 59-diphosphate-glucuronosyltransferase (UGT) isoforms (UGT1A1, 1A3, 1A4, 1A6, 1A7, 1A8, 1A9, 1A10, 2B4, 2B7, 2B10, 2B15, and 2B17) in human liver, intestinal, and kidney microsomes, and in recombinant human UGT-expressing insect cell membranes (rhUGTs) by targeted peptide–based quantification using liquid chromatography–ta...

Longstanding mechanistic questions about the role of protecting osmolyte trimethylamine N oxide (TMAO) which favors protein folding and the denaturing osmolyte urea are addressed by studying their effects on the folding of uncharged polymer chains. Using atomistic molecular dynamics simulations, we show that 1-M TMAO and 7-M urea solutions act dramatically differently on these model polymer cha...

Leuconostoc mesenteroides NRRL B-1149 produces extracellular dextransucrase which in this study was purified using different concentrations of polyethylene glycol (PEG). The dextran produced by this enzyme is unique in that it contains α(16) and α-(13) linkages which have clinical applications. The cell free supernatant with 0.9 U/mg enzyme specific activity was subjected to fractionation by ...

Horse ferricytochrome c (cyt c) undergoes exchange of one of its axial heme ligands (Met-80) for one or more non-native ligands under denaturing conditions. We have used (1)H NMR spectroscopy to detect two conformations of paramagnetic cyt c with non-native heme ligation through a range of urea concentrations. One non-native form is an equilibrium unfolding intermediate observed under partially...