George W. Rushizky

Studies on the mechanism of action of micrococcal nuclease I. Degradation of thymus deoxyribonucleic acid

Abstract 1. 1. A procedure involving paper electrophoresis and paper chromatography has been adapted for use in the resolution of products resulting from the action of micrococcal nuclease on calf-thymus DNA. 2. 2. Using this procedure, 27 mono-, di-, and trinucleotides have been isolated from the enzymic digest and identified. 3. 3. With both native and heat denatured DNA, micrococcal nuclease has been found to hydrolyze preferentially those phosphodiester bonds whose cleavage results in the liberation of fragments terminating in deoxyadenylic or thymidylic acid residues bearing free 5′-hydroxyl groups. The possibility is considered that the specificity may be related to a lower order of structural organization resulting from sequences rich in deoxyadenylic acid and thymidylic acid residues which permits better access to the enzyme. 4. 4. Certain trinucleotides, particularly those rich in guanylic acid, accumulate near the end of the enzymic reaction. Their further hydrolysis has been found tobe slow compared with the initial hydrolysis of the entire DNA molecule. 5. 5. The specificity of micrococcal nuclease enables it to be of some, although limited, use in the determination of base sequences in DNA.

Inactivation of enzyme formation by ultraviolet light. I. Action spectra and size of the sensitive unit.

Abstract The action spectra for loss of ability to form the inducible enzymes β-galactosidase and tryptophanase by Escherichia coli , and also for colony formation, resemble the absorption spectra of nucleic acids. A minimum size of the sensitive unit for enzyme formation of 300,000 molecular weight units is computed. This suggests that the sensitive unit is either high molecular weight RNA or DNA and not soluble RNA. On the basis of previously reported quantum yields, the size of the target is estimated to be somewhat larger than the minimum, perhaps of mass 700,000. This result is in fair agreement with present estimates of the size of a functional gene or of the RNA of a ribonucleoprotein particle. These data do not permit a choice between RNA and DNA as target materials.

Studies on the mechanism of action of micrococcal nuclease II. Degradation of ribonucleic acid from tobacco mosaic virus

Abstract A two-dimensional procedure employing paper electrophoresis and paper chromatography was used for the analysis of micrococcal nuclease digests of RNA from tobacco mosaic virus. The course of enzymic hydrolysis was thus followed by the quantitative comparison of the 4 mono- and 31 major oligonucleotides found in partial and exhaustive digests. The results indicate that the enzyme preferentially hydrolyzes bonds next to adenylic or uridylic acid, and is especially effective in attacking those regions of an RNA molecule in which several adenylic acid and/or uridylic acid residues are found in sequence.

Products obtained by digestion of the nucleic acids of some strains of tobacco mosaic virus with ribonuclease T1

Abstract The nucleic acids of three strains of tobacco mosaic virus were digested with RNAase T1 and the resulting products were separated by a two-dimensional mapping procedure and by paper chromatography. Maps of the RNA digests from the three strains were qualitatively very similar. Quantitative analysis of the various fractions showed no significant differences between strains M and TMV, but revealed several striking differences between strain HR and the other two strains.