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H Werbin

Publications and source records attributed to H Werbin.

At least 37 records · Page 2Linked to original sources
PubMed1977

DNA photolyase.

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Animals↗
PubMed1976

Purification from baker's yeast of an activator of DNA photolyase.

The activity of purified DNA photolyase from Baker's yeast is enhanced by a compound (Activator (III)) obtained from yeast by chloroform extraction ion exchange chromatography and gel filtration. Thin layer chromatography and spectral data indicate that the compound is homogeneous. Activator III emits at 350 and 440 nm when excited at 290 nm, and emits at 440 nm when excited at 358 nm. After acid hydrolysis, emission at 440 nm is produced only by excitation at 358 nm, indicating that activator (III) contains two separate chromophoric moieties. The chromophore excited by 358 nm light has a pK of 9-11, while the other chromophore has a pK of 4-5, and possibly of 9-11. The enhancement of photolytic activity by activator (III) at a concentration equimolar with that of the enzyme and the similarity of the fluorescent spectra of the activator with that of heat-denatured photolyase, suggests that the activator may be the chromophore associated with the enzyme.

Deoxyribodipyrimidine Photo-Lyase↗
PubMed1975

Yeast DNA photolyase: molecular weight, subunit structure, and reconstruction of active enzyme from its subunits.

Yeast DNA photolyase, purified by affinity chromatography, ran as a single component when analyzed by either electrophoresis on polyacrylamide gradient gels or by sedimentation velocity through 5-20% sucrose gradients containing 0.4 M KCl, and, therefore, was considered homogeneous. The molecular weights of photolyase, determined by these methods, were 130000 and 136000, respectively. When the enzyme was examined by electrophoresis on sodium dodecyl sulfate polyacrylamide gradient gels, it dissociated into two bands whole molecular weights were 60000 and 85000. After the enzyme was sedimented through sucrose gradients in the presence of 1.0 M KCl, two absorbance maxima, which corresponded to polypeptides of 54000 and 82500, were found in the fractions collected. Thus, the enzyme consists of two dissimilar subunits. When the two fractions that exhibited maximal absorbance were mixed together, a time-dependent increase in activity occurred, demonstrating that active enzyme could be reconstituted from these subunits. Analysis of sucrose gradients containing 1.0 M salt for photolyase activity showed that it was present exclusively in the region of the gradient corresponding to 68200 in agreement with a previous report (J. Cook and T. Worthy (1972), Biochemistry 11, 388). These active fractions were found in the overlap region between the two subunits, and their activity was attributed to reconstitution of the enzyme during the assay.

Centrifugation, Density Gradient↗
PubMed1975

Purification of an inhibitor of DNA photolyase with fluorescent spectra similar to those of the enzyme.

1. The binding of DNA photolyase, the enzyme which uncouples cyclobutadipyrimidines in DNA upon illumination, to its substrate was enhanced by a substance isolated from acidified autolysates of baker's yeast. 2. This substance, referred to as activator, was partially purified by chloroform extraction, ion-exchange chromatography on Dowex 50 and DEAE-cellulose, and gel filtration on Sephadex G-10. Upon lyophilization and storage at -20 degrees C, it was converted to a potent inhibitor of enzyme binding to substrate. 3. The inhibitor appeared as a single band in two thin-layer chromatography systems and was detected by ultraviolet absorbance, fluorescence, and ninhydrin staining. 4. The similarity between the fluorescence spectra of the inhibitor and the enzyme suggested that the inhibitor was structurally analogous to a chromophore in the enzyme and that the activator from which the inhibitor was derived may be the active chromophore of the enzyme.

Biological Assay↗