Sara Rappoport

Biochemically active sol-gel glasses: The trapping of enzymes ☆

Abstract We describe the preparation and properties of a biochemically active sol-gel glass, obtained by trapping the enzyme alkaline phosphatase (ALP) in a polymerizing tetramethoxysilane. The immobilized purified ALP from bovine intestinal mucosa had a 30% activity yield and an improved stability to thermal deactivation compared to a solution. The composite bioactive glass was preserved in water at room temperature for two months without loosing activity. A non-Michaelis-Menten kinetics was observed. The concept of preparing bioactive materials by the sol-gel method seems to be general. Thus, other enzymes (chitinase, aspartase, β-glucosidase) were successfully trapped.

Biocatalysis by sol-gel entrapped enzymes

Abstract Attachment of enzymes to insoluble matrix is an essential step in the development of biocatalysts. Transparent xerogels containing various enzymes were obtained by mixing a solution of an enzyme with tetra-methoxy orthosilicate (TMOS) at room temperature followed by gelation and drying. Effective immobilization was usually obtained at initial pH values > 7, where there is a change in the gelation mechanism from predominant hydrolysis/condensation to predominant direct polymerization of silicate precursors. The properties of sol-gel matrix, namely, transparency, large hydrophilic surface and good chemical and thermal stability, make it an ideal material for both biocatalysts and optical sensor devices. An example of a simple optical glucose sensor is demonstrated.

[55] The chemical preparation of acetylaminoacyl-tRNA☆

Publisher Summary This chapter describes the method for the chemical preparation of Acetylaminoacyl-tRNA that is based on the reaction between aminoacyl-tRNA and N-hydroxysuccinimide ester of acetic acid. The active ester is prepared by allowing the acetic acid to react with N-hydroxysuccinimide in the presence of dicyclohexylcarbodiimide. This method was found to be a general one and has been used successfully with aliphatic carboxylic acids—for example, formic, acetic, caprylic, lauric, and palmitic acids as well as with aromatic compounds containing free carboxylic groups. It was found that when N-hydroxysuccinimide ester of [ 3 H]acetic acid was allowed to react with deacylated tRNA, the radioactivity associated with the tRNA was equivalent to one acetyl residue per 100 molecules of tRNA.

Design and Properties of Enzymes Immobilized in Sol-Gel Glass Matrices

Technical convenience and the urge to lower costs are driving industrial interest, and thus applied research, toward ever-increasing effort to prepare successfully immobilized enzymes. Immobilization allows re-use of enzymes, protects them from harsh external conditions, from microbial contamination, and prolongs their useful lifetime. There are probably as many immobilization methods as there are enzymes. This proliferation of techniques reflects the complexity of the biological material and the variety of its uses. Simple inexpensive general techniques, resulting in stable and active enzyme catalysts are yet in demand (Kennedy and White, 1985a).

[56] The synthesis of oligopeptidyl-tRNA☆

Publisher Summary This chapter describes the procedures for the synthesis of oligopeptidyl-tRNA. Peptidyl-tRNA can be isolated from ribosomes active in protein synthesis. The peptidyl-tRNA so obtained is a mixture of different tRNAs to which peptides of different chain lengths and different amino acid composition are bound. By using synthetic homopolynucleotides or copolynucleotides with a known sequence, one can control the amino acid composition of the peptidyl-tRNA, but it is very difficult to control the peptide chain length. To obtain chemically defined peptidyl-tRNA in sufficient quantities for biochemical and physical investigations, peptidyl-tRNAs have been prepared chemically, using aminoacyl-tRNA as the starting material. The chemical synthesis of peptidyl-tRNA using aminoacyl-tRNA as starting material can be accomplished in three major steps: (1) preparation of a suitable N-blocked carboxyl activated amino acid or peptide, (2) condensing the N-blocked carboxyl activated amino acid (or peptide) with aminoacyl-tRNA, and (3) removing the N-blocking group from the peptidyl-tRNA.