Magnetic bead cellulose as a suitable support for immobilization of α-chymotrypsin.
Magnetic bead cellulose was prepared by a suspension method from the mixture of viscose and magnetite using thermal sol-gel transition and regeneration of cellulose. The prepared magnetic particles after their activation with divinyl sulfone were shown to be suitable magnetic carrier for immobilization of α-chymotrypsin and for its application in proteomic studies. The specific activity of the immobilized proteinase was high; its activity did not change in the course of storage. The following properties of the immobilized proteinase were compared with those of the soluble enzyme: pH and temperature dependence of the activity, self-cleavage activity, and possibility of repeated use. α-Chymotrypsin immobilized to magnetic bead cellulose was used for the proteolytic digestion of porcine pepsin A and human gastric juice and a possibility of direct use of enzyme reaction products for matrix-assisted laser desorption/ionization time of flight mass spectrometry analysis was shown.
Institute of Pathological Physiology, First Faculty of Medicine, Charles University in Prague, U Nemocnice 5, 128 53 Prague 2, Czech Republic. petr.prikryl@lf1.cuni.cz
Petr
P
Prikryl
Jiri
J
Lenfeld
Daniel
D
Horak
Marie
M
Ticha
Zdenka
Z
Kucerova
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002482:Cellulose / Q000737:chemistry*; D002918:Chymotrypsin / Q000737:chemistry* / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism; D005750:Gastric Juice / Q000201:enzymology; D006801:Humans; D059346:Magnets / Q000737:chemistry*; D008863:Microspheres; D008969:Molecular Sequence Data; D010434:Pepsin A / Q000737:chemistry / Q000378:metabolism; D059748:Proteolysis; D012995:Solubility; D013450:Sulfones / Q000737:chemistry
D004800:Enzymes, Immobilized; D013450:Sulfones; C009873:divinyl sulfone; D002482:Cellulose; D002918:Chymotrypsin; C117608:alpha-chymotrypsin; D010434:Pepsin A