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Nucleic Acids Research, 1983, Vol. 11, No. 3 659-669
© 1983

MOLECULAR BIOLOGY

A versatile method for the coupling of protein to DNA: synthesls of {alpha}{gamma} Marcroglobulin-Dna conjugates

Sheue-Yann Cheng, Glenn T. Merlino and H. Pastan

National Cancer Institute, Laboratory of Molecular Biology Bethesda, MD 20205, USA

Received December 10, 1982. Accepted January 12, 1983.

We describe a simple, general method to link proteins covalently to ONA. The method uses two reagents, N-acetyl-N'-(p-glyoxylylbenzoyl)cystam1ne and 2-1m1noth1olane. The former reacts specifically with nonpaired quanine residues and upon reduction generates a free sulfhydryl group. The latter reacts with a protein to provide another sulfhydryl group which 1s subsequently conjugated to DNA by an intermolecular disulfide Interchange reaction. Using this method {alpha}2- macroglobulin was conjugated to plasmid DNA encoding the Herpes simplex virus-1 thymidine kinase gene or a DNA fragment containing the E.coli chlor-amphenicol acetyltransferase gene. Up to 20% of the total DNA was conjugated to {alpha}2-macroglobul1n and the {alpha}2-macroglobulin-DNA conjugate had a prote1n/DNA molar ratio of approximately two. The whole reaction takes place under very mild conditions 1n aqueous solution. The structure of DNA appears not to be significantly affected by the chemical modification. This method may prove useful In ligand directed gene transfer studies.


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