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Affinity chromatography on immobilized Protein A is the current method of choice for the purification of monoclonal antibodies (mAbs). Despite its widespread use it presents certain drawbacks, such as ligand instability, leaching, toxicity and high cost. In the present work, we report a new procedure for the purification of two human monoclonal anti-HIV(More)
Tetracycline complexation with calcium and organic ligands was studied using fluorescence, circular dichroism, and solvent extraction methods. The results were used to interpret the mechanism of the commonly used fluorometric methods for the analysis of tetracycline in biological fluids. Tetracycline formed ternary calcium complexes with barbital sodium and(More)
A simple and accurate assay for the determination of griseofulvin and its metabolites in biological fluids using high-performance liquid chromatography is described. Using a reversed phase column and a mobile phase solvent of 45% acetonitrile in 0.1 M acetic acid, baseline separation of griseofulvin and several analogues was obtained. The described method(More)
Solution differential scanning calorimetry (DSC) of oxidized amicyanin, a Type I copper protein, at pH 7.5 reveals two thermal transitions. The major transition at 67.7 degrees C corresponds to the disruption of the Cys(92) thiolate to Cu(II) charge transfer as evidenced by a corresponding temperature-dependent loss of amicyanin visible absorbance. A minor(More)
Immunoreactive proteins of 45-kD and 29/30-kD doublet bands are candidate imidazoline receptor binding proteins (IRBP) based on associations with I(1) or I(2) binding sites, respectively. It was reported that the density of cortical membrane 29/30-kD I(2) protein is diminished whereas a 45-kD I(1) protein is increased in depressed suicide victims versus(More)
M98Q amicyanin is isolated with zinc bound to its type 1 copper-binding site. The influence of the axial ligand of the type 1 copper site on metal specificity is strongest prior to the completion of protein folding and adoption of the final type 1 site geometry. The preference for zinc over copper correlated with the selectivity of apoamicyanin in vitro in(More)
Amicyanin from Paracoccus denitrificans is a type 1 copper protein with three strong equatorial copper ligands provided by nitrogens of His53 and His95 and the sulfur of Cys92, with an additional weak axial ligand provided by the sulfur of Met98. Met98 was replaced with either Gln or Ala. As isolated, the M98A and M98Q mutant proteins contain zinc in the(More)
Amicyanin is a type 1 copper protein that is the natural electron acceptor for the quinoprotein methylamine dehydrogenase (MADH). A P52G amicyanin mutation increased the Kd for complex formation and caused the normally true electron transfer (ET) reaction from O-quinol MADH to amicyanin to become a gated ET reaction (Ma, J. K., Carrell, C. J., Mathews, F.(More)
Mutation of the axial Met ligand of the type 1 copper site of amicyanin to Ala or Gln yielded M98A amicyanin, which exhibits typical axial type 1 ligation geometry but with a water molecule providing the axial ligand, and M98Q amicyanin, which exhibits significant rhombic distortion of the type 1 site (Carrell, C. J., Ma, J. K., Antholine, W. E., Hosler, J.(More)