Effect of water-soluble phospholipid polymers conjugated with papain on the enzymatic stability.
To maintain enzymatic activity during long-term storage by conjugation with water-soluble 2-methacryloyloxyethyl phosphorylcholine (MPC) polymers (PMPC-COOH) having various molecular weights with a carboxylic group on the terminal, such compounds were synthesized as a polymer modifier using a photoinduced living radical polymerization technique. A poly(ethylene oxide) with a carboxyl group (PEO-COOH) was used as the control. The PMPC-COOHs were reacted with the amino groups of the enzyme, papain, via amide bonds. With an increase in the molecular weight in the range between 5 and 20K of the PMPC-COOH, the modification degree and alpha-helix content of the conjugated papain slightly decreased, but the remaining enzymatic activity did not depend on the molecular weight of the PMPC-COOH. However, when a much higher molecular weight PMPC-COOH (40K) was conjugated with a reduction in the modification degree, alpha-helix content was higher compared with the other PMPC-conjugated papain. Modification with PEO-COOH showed little reduction of the alpha-helix content of papain. The time dependence of the remaining enzymatic activity of the polymer-conjugated papains was evaluated during storage at 40 degrees C. The native papain diminished activity within one week. PEO-conjugated papain had decreased activity with time, but after one week it had half its initial level. The same tendency was observed when papain was modified with PMPC-COOHs 5 and 40K, that is, the enzymatic activity did not decrease even when they were stored for 4 weeks. We concluded that the PMPC chain could stabilize the enzyme by control of the molecular weight of the PMPC and modification degree to the enzyme.
Miyamoto
Daisuke
D
Department of Materials Engineering, School of Engineering, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, 113-8656, Tokyo, Japan.
Watanabe
Junji
J
Ishihara
Kazuhiko
K
D001672:Biocompatible Materials / Q000138:chemical synthesis / Q000737:chemistry*; D004789:Enzyme Activation; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry; D008422:Materials Testing; D008689:Methacrylates / Q000737:chemistry*; D010206:Papain / Q000737:chemistry*; D010743:Phospholipids / Q000737:chemistry*; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry; D011109:Polymethacrylic Acids; D011485:Protein Binding; D012995:Solubility; D014867:Water / Q000737:chemistry*
D001672:Biocompatible Materials; D004800:Enzymes, Immobilized; D008689:Methacrylates; D010743:Phospholipids; D011108:Polymers; D011109:Polymethacrylic Acids; C115766:poly(2-methacryloyloxyethyl-phosphorylcholine); D014867:Water; D010767:Phosphorylcholine; D010206:Papain