Tag Archives: gel-filtration

Properties of alpha-L-fucosidase for serum of patients with hepatocellular cancer and cytotoxicity on some cancer cell lines

Z. M. A. A. Hamodat

University of Mosul, College of Science, Chemistry Department, Mosul – Iraq;
e-mail: zahraahamodat@uomosul.edu.iq

Received: 07 July 2021; Accepted: 12 November 2021

Alpha-L-fucosidase (FUCA) degrades many fucosylated glycans and has long been recognized as a tumor marker associated with the early detection of some cancers. This study aimed to purify and characterize alpha-L-fucosidase from the serum of patients with hepatocellular cancer and estimate its toxic effect against hepatocellular carcinoma HepG2, prostate cancer PC3 cell lines and the standard hepatocyte WRL-68 cell line. SDS-Page Electrophoresis technique was used to determine the purity of the purified alpha-L fucosidase and estimate its molecular weight. Three purification steps were used for FUCA purification: precipitation with 65% ammonium sulfate saturation, DEAE-cellulose ion exchange, Sephadex G-75 gel filtration. The procedure resulted in 54% recovery of the enzyme with 27.5-fold purification and 14 U/mg specific activity. It was demonstrated that FUCA purified from the serum of HCC patients showed a more toxic effect on HepG2 cells (IC50 of 65.74 µg/ml) than on PC3 prostate cancer cells (IC50 of 111.5 g/ml) and less toxic effect against standard hepatocyte WRL-68cells (IC50 of 214.5 µg/ml). We can conclude that the inhibitory effect of the purified FUCA on hepatocellular carcinoma is more than its effect on prostate cancer cells. Also, the purified FUCA may be used in studies on anticancer drug development in liver cancer.

Isolation of κ-CN-1P and β-CN-5Р fractions from native casein micelles

V. G. Yukalo, L. A. Storozh

Ternopil Ivan Puluj National Technical University, Ukraine;
e-mail: biotech@tu.edu.te.uaβ

Proteins of the casein complex of milk arouse considerable interest as the precursors of biologically active peptides which are capable of influencing various physiological systems of the body (digestive, nervous, cardiovascular, and immune). It has been established that various bioactive peptides are unevenly located in the structure of casein fractions. In this connection, there appeared a need to separate individual fractions of this protein for studying the pathways of formation and properties of bioactive casein peptides. To minimize negative effects of the purification procedure, we used the gel filtration on Sefarose 2B to produce native casein micelles and repeating filtration on Sephadex G-150 to separate individual fractions. As a result, according to electrophoretic analysis, casein micelles with a characteristic protein composition were obtained. Taking into account the similarity of the molecular weight of components for their dividing the repeated gel filtration was carried out with separating the chromatograms into sectors. The composition of the combined fractions of each sector was analyzed by electrophoresis. This approach allowed isolating two electrophoretically homogeneous proteins from native casein micelles – κ-CN-1P and β-CN-5P, as well as to obtain a substantially purified (> 83%) αS1-CN-XP. Isolated caseins without the influence of extreme factors can be used to study natural bioactive peptides.


RNP-complex stoichiometry of Bombyx mori nuclear polyhedrosis virus polyhedra

T. V. Shirina, M. T. Bobrovskaja, E. A. Kоzlov

Institute of Molecular Biology and Genetics,
National Academy of Sciences of Ukraine, Kyiv;
e-mail: e.a.kozlov@imbg.org.ua

By gel-filtration through Sephacryl S-300 it was shown that RNP A complex present in polyhedra of Bombyx mori nuclear polyhedrosis virus has molecular weight (Mw) about 700 kDa. It was shown that RNP A with Mw 788 kDa is composed of two polyhedrin 13S-associates with Mw 342 kDa, two p14 polypeptide with Mw 14 kDa, two 21 kDa small non-coded RNAs and two 17 kDa small non-coded RNAs. The model of RNP A formation from components making it is proposed. The complex role in the course of polyhedron formation and its role in the course of infection are discussed.