Methods for preparing proteins of altered structure

Details Methods for preparing proteins of altered structure Methods for preparing proteins of altered structure

C - Chemistry, Metallurgy

12

N

195/1.12, 195/1.

C12N 15/11 (2006.01) C07H 21/00 (2006.01) C12N 15/00 (2006.01) C12N 15/10 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 1323318

ABSTRACT The present invention relates to methods for inducing cellular mutation by promoting the reaction of nonenzymatic glycosylation of genetic material, such as DNA and RNA, by incubation of the genetic material with certain protein structure modifying agents. Suitable such agents are reducing sugars, such as glucose, glucose-6-phosphate and a reactive intermediate comprising a complex between glucose 6-phosphate and lysine. The methods include the in vitro reaction of the agent with genetic material, such as a particular gene-bearing plasmid, under conditions promoting mutation followed by the introduction of the plasmid bearing the reacted gene into a host cell, and allowing the reacted gene to be mutated and expressed, after which the mutant protein of interest may be isolated. In a variation on the last-mentioned method, a first genetic material may be incubated with the agent and reacted, and this reacted genetic material may then be introduced into a distinct cellular strain and appropriately incubated whereupon the first reacted genetic material will cause the mutation of the genetic material in the distinct cellular strain. The invention also includes an in vivo embodiment wherein the particular genetic material of interest may be isolated and introduced into a host cell which contains high intracellular concentrations of the protein structure modifying agent(s). In this instance, the mutation is promoted by the host cell, and the mutant material may thereafter be isolated and recovered. Diagnostic methods employing the principles of the invention are also disclosed.

588099

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