Bioaffinity Based Immobilization of Enzymes - Advances in Biochemical Engineering Biotechnology - page 199

Bioaffinity Based Immobilization of Enzymes

M. Saleemuddin

Department of Biochemistry,Faculty of Life Sciences and Interdisciplinary Biotechnology
Unit,Aligarh Muslim University,Aligarh - 202 002,India. E-mail: [e-mail]

Procedures that utilize the affinities of biomolecules and ligands for the immobilization of
enzymes are gaining increasing acceptance in the construction of sensitive enzyme-based
analytical devices as well as for other applications. The strong affinity of polyclonal/mono-
clonal antibodies for specific enzymes and those of lectins for glycoenzymes bearing appro-
priate oligosaccharides have been generally employed for the purpose. Potential of affinity
pairs like cellulose-cellulose binding domain bearing enzymes and immobilized metal ion-
surface histidine bearing enzymes has also been recognised. The bioaffinity based immobiliza-
tion procedures usually yield preparations exhibiting high catalytic activity and improved
stability against denaturation. Bioaffinity based immobilizations are usually reversible facili-
tating the reuse of support matrix,orient the enzymes favourably and offer the possibility of
enzyme immobilization directly from partially pure enzyme preparations or even cell lysates.
Enzyme lacking innate ability to bind to various affinity supports can be made to bind to them
by chemically or genetically linking the enzymes with appropriate polypeptides/domains like
the cellulose binding domain,protein A,histidine-rich peptides,single chain antibodies,etc.

Keywords: Concanavalin A,Monoclonal Antibodies,Polyclonal Antibodies,Reloadable Bio-
sensors,Antibody Orientation,Fusion Proteins,Glycoenzymes,Immobilized Metal Ion Sup-
ports,Enzyme Stabilization.

1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204

2 Immunoaffinity Immobilization of Enzymes . . . . . . . . . . . 205

2.1 Monoclonal or Polyclonal Antibodies? . . . . . . . . . . . . . . . 206

2.2 Selection of the Antibodies . . . . . . . . . . . . . . . . . . . . . . 208

2.3 Immunoaffinity Immobilization Strategies . . . . . . . . . . . . . 209

2.3.1 Immobilization without Solid Supports . . . . . . . . . . . . . . . 209

2.3.2 Construction of Immunoadsorbents . . . . . . . . . . . . . . . . 209

2.3.2.1 Favourable Orientation of Antibodies on Supports . . . . . . . . 210

2.3.3 Use of Secondary Antibodies . . . . . . . . . . . . . . . . . . . . . 211

2.4 Reusability of Immunoaffinity Supports . . . . . . . . . . . . . . 212

2.5 Behaviour of Immunoaffinity Immobilized Enzymes . . . . . . . 213

2.6 Utility in Organic Solvents . . . . . . . . . . . . . . . . . . . . . . 214

3 Lectin Affinity Based Immobilization of Glycoenzymes . . . . . 214

3.1 Concanavalin A . . . . . . . . . . . . . . . . . . . . . . . . . . . . 214

3.2 Immobilization of Glycoenzymes Using Concanavalin A . . . . . 215

3.3 Immobilization ofWhole Cells . . . . . . . . . . . . . . . . . . . 218

3.4 Immobilization with Other Lectins . . . . . . . . . . . . . . . . . 218