Behavior of the human a1-acid glycoprotein in water–low aliphatic alcohol systems
L. Maršálková1, V. Kopecký Jr.2, V. Karpenko1
1 Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University, Albertov 6, CZ-12843 Prague 2, Czech Republic
2 Institute of Physics, Faculty of Mathematics and Physics, Charles University, Ke Karlovu 5, CZ-12116 Prague 2, Czech Republic; firstname.lastname@example.org
a1-acid glycoprotein (AGP), also known as orosomucoid, is a 41-kDa single polypeptide formed of 183 amino acids with 21 possible substitutions in one polypeptide chain linked by two disulfide bridges. Carbohydrate moiety is slightly over 40% of its total molecular weight. It is formed of five branched carbohydrate units linked to Asp of the peptide chain by the N-glycosidic bond and terminated by sialic acid residues . Concerning useful chromophore groups there are 3 Trp, 12 Tyr, and up to 12 Phe residues in AGP molecule. The native structure of AGP is characterized by low a-helix content and is dominated by b-sheets . A model of three-dimensional structure of AGP has been published recently , which have been proposed by an approach that combines molecular modeling and vibrational spectroscopy. It shows that AGP folds as a highly symmetrical all-b protein dominated by a single eight-stranded antiparallel b-sheet. AGP is a glycoprotein that occurs physiologically in human blood serum. It is known that AGP plays a role under inflammatory or other pathological conditions and is able to bind basic drugs and certain steroid hormones such as progesterone. However, its biological function and complete 3D structure remains unknown.
AGP has been studied over the whole pH range in the aqueous solutions. Behavior in the presence of alcohol has been examined only in the basic region . We focused our attention on acid pH range, because isoelectric point of AGP varies between 1.9 and 2.7 depending on the buffer used for its determination. It is supposed that presence of alcohol in the system influences both intra- and intermolecular interactions of this protein. Alcohols may cause changes in the structure of water and, consequently, the hydration shell of AGP molecule is likely to be disturbed. Furthermore, alcohols decrease permittivity of the solvent, and thus the electrostatic interactions in the system.
The aim of the present work was to examine the effect of four aliphatic alcohols – methanol, ethanol, 1‑propanol and 2‑propanol, on the structure of AGP molecule in the acid region. Weight fraction of all alcohols was 0.10; the experiments have been performed in the pH region 2–7. Behavior of AGP in the presence of alcohols has been studied by means of infrared spectroscopy, UV-VIS absorption spectroscopy and fluorescence spectroscopy. Spectral sets of each used spectroscopic method used (at desired pH or in presence of an aliphatic alcohol) have been subsequently examined by principal component analysis. The two-dimensional (2D) correlation spectroscopy was applied as well as a new method of a multivariate statistics which provides the specific order of the spectral intensity changes taking place during the measurement on the value of variable affecting the spectra. Moreover, heterospectral 2D correlation spectroscopy enables joint data evaluations of spectral sets measured by different spectroscopic methods at the same conditions. Finally, it has been demonstrated that the presence of alcohols in the most acid part of the studied pH region has significantly influenced AGP structure, particularly it has led to decrease of b‑sheet content.
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Ministry of Education of the Czech Republic and the Grant Agency of the Czech Republic are gratefully acknowledged for support (No. MSM 0021620835, No. 202/06/P208, respectively).