Biomaterial Database

5'-Nucleotidase: solubilization, radiochemical analysis, and electrophoresis. 1979

C Tucker-Pian, and B Bakay, and W L Nyhan

5'-Nucleotidase (5'-NT, E.C. 3.1.3.5) of cultured human and rodent cells was rendered soluble using the zwitterionic detergent Zwittergent 314. Optimal activity of 5'-NT was obtained when sonicated cells were incubated in solutions containing 0.75% (w/v) Zwittergent. A method was developed for the determination of the activity of 5'-NT in which the unutilized substrate, [14C]-AMP, was precipitated with lanthanum chloride and the soluble [14C]-adenosine was measured by scintillation counting. 5'-NT isozymes were separated using agarose gel electrophoresis and isoelectric focusing in polyacrylamide gel. The zones of enzyme activity were established by precipitation of unutilized [14C]-AMP with LaCl3, removal of soluble [14C)-adenosine by washing gels in water, and autoradiography. The zones of 5'-NT appeared as clear zones on darkened X-ray film. When analyzed by agarose gel electrophoresis, fibroblasts derived from human skin and rat liver produced a single zone of 5'-NT activity. The 5'-NT isozyme of rat cells migrated faster than that of human cells and was easy to distinguish. The presence of detergent in the sample and in the gel enhanced enzymatic activity and improved the separation of the isozymes. Isoelectric focusing resolved 5'-NT of human fibroblasts into two molecular forms, one of which focused in the region of pH 6 and the other at pH5.

UI	MeSH Term	Description	Entries
D007525	Isoelectric Focusing	Electrophoresis in which a pH gradient is established in a gel medium and proteins migrate until they reach the site (or focus) at which the pH is equal to their isoelectric point.	Electrofocusing,Focusing, Isoelectric
D008099	Liver	A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.	Livers
D009708	Nucleotidases	A class of enzymes that catalyze the conversion of a nucleotide and water to a nucleoside and orthophosphate. EC 3.1.3.-.
D002460	Cell Line	Established cell cultures that have the potential to propagate indefinitely.	Cell Lines,Line, Cell,Lines, Cell
D002462	Cell Membrane	The lipid- and protein-containing, selectively permeable membrane that surrounds the cytoplasm in prokaryotic and eukaryotic cells.	Plasma Membrane,Cytoplasmic Membrane,Cell Membranes,Cytoplasmic Membranes,Membrane, Cell,Membrane, Cytoplasmic,Membrane, Plasma,Membranes, Cell,Membranes, Cytoplasmic,Membranes, Plasma,Plasma Membranes
D003902	Detergents	Purifying or cleansing agents, usually salts of long-chain aliphatic bases or acids, that exert cleansing (oil-dissolving) and antimicrobial effects through a surface action that depends on possessing both hydrophilic and hydrophobic properties.	Cleansing Agents,Detergent Pods,Laundry Detergent Pods,Laundry Pods,Syndet,Synthetic Detergent,Agent, Cleansing,Agents, Cleansing,Cleansing Agent,Detergent,Detergent Pod,Detergent Pod, Laundry,Detergent Pods, Laundry,Detergent, Synthetic,Detergents, Synthetic,Laundry Detergent Pod,Laundry Pod,Pod, Detergent,Pod, Laundry,Pod, Laundry Detergent,Pods, Detergent,Pods, Laundry,Pods, Laundry Detergent,Synthetic Detergents
D004587	Electrophoresis, Agar Gel	Electrophoresis in which agar or agarose gel is used as the diffusion medium.	Electrophoresis, Agarose Gel,Agar Gel Electrophoresis,Agarose Gel Electrophoresis,Gel Electrophoresis, Agar,Gel Electrophoresis, Agarose
D006224	Cricetinae	A subfamily in the family MURIDAE, comprising the hamsters. Four of the more common genera are Cricetus, CRICETULUS; MESOCRICETUS; and PHODOPUS.	Cricetus,Hamsters,Hamster
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia