Document Type : Research Paper
Authors
University Of Baghdad - College of Science
Abstract
The aim of this work is to characterize spectrophotometrically the isolated Alpha fetoprotein from human colorectal tumor homogenates and the molecules of both AFP antibody and the complex of AFP/anti AFP antibody.Gel filtration technique was used to separate 125I-anti AFP antibody bound to human AFP from unbound (free) 125I-anti AFP antibody. The characterization of human-AFP, anti-AFP, and (AFP/Anti-AFP) complex were carried out through the ultraviolet (U.V) spectroscopic studies.Factors affecting the light absorption properties of the molecules under investigation in this work such as pH, solvent polarity (solvent perturbation technique), spectrophotometric pH titration and thermal stability have been studied.The spectrophotometric pH titration for h-AFP, anti AFP, and (AFP/anti-AFP) complex showed that pKa for tyrosine was 9.5, 10.2, and 9.9, while for histidine was 5.7, 6.0, and 5.9 respectively. Spectrophotometric pH titration and several spectral changes were obtained in the presence of different polar and non-polar solvents, like the alteration of max position and intensities of protein spectrum, and the appearance of new chromophores on the surface of protein molecule. These chromophores where embedded in an interior region of the protein in the absence of the solvent.The difference in pH and polarity of the solvents is very important thing to characterize the protein molecules spectrophotometrially because they change the positions and values of molecules λmax in the UV region.
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