Polyacrylamide gels copolymerized with active esters. A new medium for affinity systems.

A new and versatile method for linking biologically active ligands to a polyacrylamide matrix is reported. Active esters of acrylic acid (N-succinimicyl acrylate and N-phthalimidyl acrylate) were synthesized, then copolymerized with acrylamide and N,N'-methylenebisacrylamide. Displacement of the active ester in the gel thus formed by various ligands containing aliphatic amino groups resulted in the formation of stable amid bonds between the ligands and the polyacrylamide gel. The affinity gel thus prepared has the following advantages: (i) resistance to chemical and microbiological degradation, (ii) ease of control of ligand level and higher levels of ligand possible, (iii) ease of control of porosity, and (iv) total displacement of the active ester under suitable conditions. Efficacy of this system was tested by preparation of 6-aminohexyl 2-acetamido-2-deoxy-beta-D-glucopyranoside derivative polyacrylamide gel by the described method. It was found to be more effective for purification of wheat germ agglutinin than the previously published affinity chromatography systems and the wheat germ hemagglutinin was obtained in crystalline form. In addition, partial resolution of isolectins was obtained from the affinity gel witha pH gradient.

[1]  M. Burger,et al.  Wheat germ agglutinin. Molecular characteristics and specificity for sugar binding. , 1974, The Journal of biological chemistry.

[2]  M. Burger,et al.  Purification of wheat germ agglutinin using affinity chromatography on chitin. , 1974, Biochemical and biophysical research communications.

[3]  V. Hořejší,et al.  Studies on phytohemagglutinins XVIII. Affinity electrophoresis of phytohemagglutinins , 1974 .

[4]  C. L. Yuan,et al.  Synthesis of 1-thioaldosides having an amino group at the aglycon terminal , 1973 .

[5]  R. Hill,et al.  Purification of wheat germ agglutinin by affinity chromatography. , 1973, Analytical biochemistry.

[6]  N. Sharon,et al.  The biochemistry of plant lectins (phytohemagglutinins). , 1973, Annual review of biochemistry.

[7]  A. Neuberger,et al.  The purification, composition and specificity of wheat-germ agglutinin. , 1973, The Biochemical journal.

[8]  R. Hill,et al.  Agarose derivatives of uridine diphosphate and N-acetylglucosamine for the purification of a galactosyltransferase. , 1972, The Journal of biological chemistry.

[9]  P. Cuatrecasas,et al.  Adsorbents for affinity chromatography. Use of N-hydroxysuccinimide esters of agarose. , 1972, Biochemistry.

[10]  M. Burger,et al.  Wheat germ agglutinin. Isolation and crystallization. , 1972, The Journal of biological chemistry.

[11]  Y. C. Lee.  [6] Analysis of sugars by automated liquid chromatography , 1972 .

[12]  V. Marchesi.  [42] Wheat germ (Triticum vulgaris) agglutinin , 1972 .

[13]  P. Cuatrecasas,et al.  The purification of beta-galactosidase from Escherichia coli by affinity chromatography. , 1971, The Journal of biological chemistry.

[14]  P. Cuatrecasas,et al.  Protein purification by affinity chromatography. Derivatizations of agarose and polyacrylamide beads. , 1970, The Journal of biological chemistry.

[15]  H. Dintzis,et al.  The derivatization of cross-linked polyacrylamide beads. Controlled introduction of functional groups for the preparation of special-purpose, biochemical adsorbents. , 1969, Biochemistry.

[16]  Yuan-chuan Lee,et al.  Microheterogeneity of the carbohydrate group of Aspergillus oryzae α-amylase , 1969 .

[17]  M. Burger,et al.  Identification of a tumor-specific determinant on neoplastic cell surfaces. , 1967, Proceedings of the National Academy of Sciences of the United States of America.

[18]  I. Silman,et al.  Water-insoluble derivatives of enzymes, antigens, and antibodies. , 1966, Annual review of biochemistry.

[19]  G. W. Anderson,et al.  The Use of Esters of N-Hydroxysuccinimide in Peptide Synthesis , 1964 .

[20]  G. I. Tesser,et al.  A NOVEL ACTIVATED ESTER IN PEPTIDE SYNTHESES , 1961 .

[21]  O. H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[22]  F. Lipmann,et al.  A SPECIFIC MICROMETHOD FOR THE DETERMINATION OF ACYL PHOSPHATES , 1945 .