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Photoreactivating enzyme from Escherichia coli: isolated enzyme lacks absorption in its actinic wavelength region and its ribonucleic acid cofactor is partially double stranded when associated with apoprotein
- Source :
- Biochemistry. 21:3914-3921
- Publication Year :
- 1982
- Publisher :
- American Chemical Society (ACS), 1982.
-
Abstract
- Isolated photoreactivating enzyme (PRE) from Escherichia coli exhibits some optical density at wavelengths greater than 300 nm. After correcting for the effects of light scattering, however, we find no true absorption in the spectral region that is required for enzymatic activity (320-450 nm). At shorter wavelengths, there is an absorption maximum near 260 nm that is due primarily to an RNA cofactor. Heating to 60/sup 0/ C and subsequently cooling to 4/sup 0/ C released the RNA cofactor from association with apoprotein and result in hyperchromicity. Circular dichroism indicates that the RNA associated with native enzyme is partially double stranded. At low ionic strength (..mu.. = 0.01), heating to 15/sup 0/ C or protease treatment at 4/sup 0/ C results in irreversible loss of part of the double strandedness. We show that the difference spectrum at 4/sup 0/ C between the absorption spectra of native enzyme and heat-treated enzyme can be fit by a superposition of reference spectra for denaturation of A-U and G-C base pairs derived from model polynucleotides. The coefficients of the linear combination of reference spectra were used to calculate the fraction of A-U and G-C base pairs. We find that both A-U and G-C base pairsmore » are present in equal concentrations and that about 20% are in a double-standard conformation in the native enzyme.« less
- Subjects :
- Circular dichroism
Hot Temperature
Absorption spectroscopy
Protein Conformation
Stereochemistry
Base pair
Lyases
Biochemistry
Cofactor
Escherichia coli
Denaturation (biochemistry)
RNA, Double-Stranded
Base Composition
biology
Chemistry
Circular Dichroism
Hyperchromicity
Dichroism
RNA, Bacterial
Pyrimidine Dimers
Spectrophotometry
Polynucleotide
biology.protein
Nucleic Acid Conformation
Deoxyribodipyrimidine Photo-Lyase
Peptide Hydrolases
Subjects
Details
- ISSN :
- 15204995 and 00062960
- Volume :
- 21
- Database :
- OpenAIRE
- Journal :
- Biochemistry
- Accession number :
- edsair.doi.dedup.....81f11b1c21d33f35c36a54600adac9f6