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Biomedical subjects

S Linn

Publications and source records attributed to S Linn.

At least 199 records · Page 11Linked to original sources
PubMed1978

Human uracil DNA N-glycosidase: studies in normal and repair defective cultured fibroblasts.

Uracil DNA N-glycosidase, an enzyme which participates in the excision of uracil from DNA, was measured in extracts from fibroblasts lines cultured from normal subjects, from several subjects with the genetic disease xeroderma pigmentosum, and from a subject with ataxia telangiectasia. The cell lines representative of complementation groups A and D of xeroderma pigmentosum and of ataxia telangiectasia had roughly the same level of activity as did the normal cells. On the other hand, cells from two xeroderma pigmentosum variants (XP4BE and XP13BE) had roughly half the normal level of activity, and cells from the heterozygous mother of XP4BE had an intermediate level of activity. In spite of these quantitative differences, no systematic alterations in reaction characteristics, apparent Km for substrate, or purification characteristics were noted for enzyme from any of the lines. Thus a causal relationship, if any, between levels of activity and the disease symptoms is equivocal.

Apurinic Acid↗
PubMed1978

Xeroderma pigmentosum fibroblasts of the D group lack an apurinic DNA endonuclease species with a low apparent Km.

Apurinic DNA endonuclease activity from cultured human fibroblasts was resolved into two species by phosphocellulose chromatography. The species had sedimentation coefficients of 3.3 S and 2.8 S and apparent Km's for apurinic sites of 5 and 44 nM, respectively. The low Km species was absent from extracts of cell lines XP5BE, XP6BE and XP7BE of xeroderma pigmentosum complementation group D.

Apurinic Acid↗
PubMed1977

Endonuclease from Escherichia coli that acts specifically upon duplex DNA damaged by ultraviolet light, osmium tetroxide, acid, or x-rays.

An endonuclease which is active upon DNA exposed to ultraviolet light at a photoproduct other than thymine dimers has been extensively purified from Escherichia coli. The small (2.7 S) enzyme is active in the presence of EDTA, has a neutral pH optimum, and is inhibited by tRNA and 1 M NaCl. It has no detectable exonuclease, DNA-N-glycosidase, or ribonuclease activities. The enzyme also nicks duplex DNA exposed to OsO4, x-rays, or acid, but it does not act upon undamaged DNA or irradiated single-stranded DNA. The majority of sites of action in DNA exposed to ultraviolet light or OsO4 appear to be alkali-stable, but those in DNA exposed to x-rays or acid are not. The incisions created by the endonuclease contain 5'-phosphate termini. The enzyme is possibly the same as E. coli endonuclease III described by Radman (Radman, M. (1976) J. Biol. Chem. 251, 1438-1445), but it is distinguishable from the other endodeoxyribonucleases described from that organism.

Bacteriophages↗
PubMed1977

Endonuclease V of Escherichia coli.

A small endodeoxyribonuclease )2.3 S) that is active on single-stranded DNA has been extensively purified from Escherichia coli so as to be free of other known DNases. It has an alkaline pH optimum (9.5), requires Mg2+, and makes 3'-hydroxy and 5'-phosphate termini. The nuclease nicks duplex DNA, particularly if treated with OsO4, irradiated with ultraviolet light, or exposed to pH 5. The uracil-containing duplex DNA from the Bacillus subtilis phage PBS-2 is an especially good substrate; it is made acid-soluble by levels of the enzyme which fail to produce any acid-soluble material in other single-stranded or duplex DNAs. Neither RNA nor RNA-DNA hybrid are degraded by the enzyme. The enzyme specificity suggests that it might act at abnormal regions in DNA, so that its in vivo function could be to initiate an excision repair sequence. Its high activity on uracil-containing DNA could imply that the enzyme provides an alternative mechanism for excising uracil residues from DNA to the pathway utilizing uracil-DNA N-glycosidase. We suggest that this enzyme be designated as endonuclease V of E. coli.

Cations, Divalent↗
PubMed1977

Human endonuclease specific for apurinic/apyrimidinic sites in DNA. Partial purification and characterization of multiple forms from placenta.

Six chromatographically distinct forms of endonuclease active on apurinic and apyrimidinic sites in DNA have been purified away from DNA phosphatases, DNA N-glycosidases, and other DNases of human placenta. The forms seem to be monomeric proteins of 27,000 to 31,000 daltons, and although catalytically similar, they can be distinguished from one another on the basis of substrate Km and the effects of small molecules such as ATP. Analysis of enzymatic activity on a spectrum of damaged DNA substrates indicates that the enzyme forms probably act at an appreciable rate only adjacent to the phosphodiester bond of a deoxyribose lacking a base (purine or pyrimidine) in duplex DNA; such sites can be formed by treating the DNA with acid, alkylating agents, DNA N-glycosidases, and, probably, x-rays and OsO4. The incision is made so as to form a deoxyribose 5'-phosphate and a 3'-hydroxynucleotide.

Adenosine Triphosphate↗
PubMed1977

Characterization of an endonuclease associated with the drug resistance plasmid pKM101.

An endonuclease was detected in strains of Salmonella typhimurium containing the drug resistance plasmid pKM101. The enzyme was not detectable in strains lacking this plasmid, but it was present in strains containing mutants of pKM101 that were no longer able to enhance host cell mutagenesis. The endonuclease had a molecular weight of roughly 75,000 and, at pH 7.0, was equally active on single-stranded and duplex deoxyribonucleic acid (DNA). The reaction with single-stranded DNA was optimal at pH 5.5, whereas with duplex DNA the optimum was pH 6.8. The enzyme required a divalent cation for activity, and it had no detectable exonuclease activity with single-stranded or duplex DNA. The endonuclease extensively degraded DNA with no apparent base specificity, forming 5'-phosphomonoester termini. Although characterization of the endonuclease has not revealed its function, the enzyme does not appear to be a restriction endonuclease.

DNA, Bacterial↗
PubMed1976

Selective inhibition of the dnase activity of the recBC enzyme by the DNA binding protein from Escherichia coli.

In the presence of the Escherichia coli DNA binding protein, single-stranded DNA is resistant to both the endo- and exonucleolytic activities of the recBC DNase. Linear duplex DNA, on the other hand, is unwound at a normal rate, but converted to large, single-stranded fragments which are resistant to further hydrolysis. Therefore, in the presence of the binding protein and linear duplex DNA, the recBC enzyme acts not as a DNase, but primarily as an ATP-dependent unwinding enzyme, able to generate large, single-stranded material. Duplex circular DNA containing short, single-stranded gaps is also resistant to the hydrolysis in the presence of the binding protein.

Adenosine Triphosphate↗
PubMed1976

An altered apurinic DNA endonuclease activity in group A and group D xeroderma pigmentosum fibroblasts.

Endonuclease activity upon depurinated DNA was measured in extracts of cultured fibroblasts from xeroderma pigmentosum patients. Cell lines from complementation groups A, B, C, E, and the XP-variant had slightly reduced levels of activity, but cell lines from complementation group D had one-sixth of the normal activity. An altered pH dependence and a higher apparent Km for substrate in D-cell lines indicate that the remaining activity is also qualitatively different from the activity found in normal cells. A higher Km was also found in cell lines from the A-complementation group but not in cell lines from the C-complementation group. These defects in apurinic DNase might account for the neurological disorders in patients from the D- and the A-complementation groups.

Apurinic Acid↗
PubMed1976

Decreased fidelity of DNA polymerase activity isolated from aging human fibroblasts.

DNA polymerase (deoxynucleosidetriphosphate: DNA nucleotidyltransferase, EC 2.7.7.7 or DNA nucleotidyltransferase) activity, isolated from late and early passage cells of the diploid human fibroblast line, MRC-5, was compared. The level of activity dropped with increasing passage. In addition, when the fidelity of polymerization was monitored with four synthetic templates under a variety of conditions, it was observed that the enzyme from late passage cells was more error-prone. The possible relation of these observations to "senescence" of the fibroblasts is discussed.

DNA Nucleotidyltransferases↗
PubMed1976

Infant responsivity to curvature.

The reactions of 112 10-month-old male infants to the property of curvature were examined using a habituation paradigm with lever pressing as an instrumental response. 4 levels of curvature (straightt line, minimal curve, moderate curve, large curve) each served as a standard (habituation) stimulus and as a transformation stimulus in 9 separate experimental conditions. Results revealed a greater initial dishabituation of reinforced instrumental responding in those conditions crossing the curved/straight boundary; furthermore, this initial dishabituation was sustained throughout the transformation phase of the experiment in those conditions which involved changes from straight to curved, but not curved to straight. Results are interpreted as implying a special attention-recruiting value for curvature in 10-month old infants.

Attention↗
PubMed1975

The gamma protein specified by bacteriophage gamma. Structure and inhibitory activity for the recBC enzyme of Escherichia coli.

The protein encoded by the gam gene of bacteriophage lambda ("gamma protein") is a specific inhibitor of the recBC enzyme of Escherichia coli. The lambda protein has been purified approximately 2,000-fold, and its structure and inhibitory activity have been characterized. It appears to be composed of two identical subunits of 16,500 daltons, inhibits all of the catalytic activities of the recBC enzyme with apparently equal efficiency, but has no effect upon any other E. coli or lambda-DNase tested. Inhibition does not occur unless recBC enzyme is exposed to gamma protein prior to reaction of the enzyme with DNA. The inhibitory activity is independent of temperature, and no catalytic activity has been detected that might fulfill the inhibitory function. It appears instead that the inhibition involves a stoichiometric, rather than a catalytic interaction between gamma protein and the enzyme. Reaction kinetics for the recBC enzyme inhibited by gamma protein show no anomalous protein--only a depressed rate. Inhibition is not competitive and does not appear to affect the enzyme's affinity for DNA. The enzyme remains inhibited after it is separated from "excess" gamma protein by gel filtration or sedimentation in a glycerol gradient, and inhibited enzyme has a reduced electrophoretic mobility compared to that of uninhibited enzyme. Gamma Protein inhibits recBC enzyme which has been reconstituted from cell-free extracts by complementation in vitro, but at least one of the complementing factors present in extracts from recB- cells does not by itself form a complex with gamma protein. The mechanism of inhibition and the implications of these results from gamma replication and recombination are discussed.

Adenosine Triphosphatases↗
PubMed1974

Molecular structure of exonuclease I from Escherichia coli B.

Exonuclease I of Escherichia coli B (EC 3.1.4.25) was determined to be a monomeric protein of molecular weight approx. 72,000, as estimated by Sephadex gel filtration, sedimentation velocity centrifugation, and sodium dodecylsulfate polyacrylamide gel electrophoresis.

Centrifugation, Density Gradient↗
PubMed1974

Partial purification and characterization of four endodeoxyribonuclease activities from Escherichia coli K-12.

Four hitherto undescribed endodeoxyribonucleases, temporarily designated A(1), A(2), A(3), and B, have been isolated from E. coli K-12. Each requires Mg(++) and is not stimulated by ATP or S-adenosylmethionine. A(3) is strongly inhibited by Fe(+++) and weakly inhibited by ATP, S-adenosylmethionine, and DPN, whereas B is inhibited by caffeine. Each can be purified free of exonuclease or DNA-3'-phosphatase. A(1) (molecular weight approximately 72,000) cleaves single-stranded, circular fd DNA to form 3'-hydroxyl termini and introduces nicks and breaks in the closed, double-stranded replicative form DNA of fd (fd RFI). A(2) (molecular weight approximately 46,000) cleaves fd DNA and introduces nicks and breaks in RFI, forming 3'-hydroxyl- and 5'-phosphoryl termini. A(3) (molecular weight approximately 38,000) cleaves fd DNA to form 3'-hydroxyl termini and introduces only nicks in fd RFI. Irradiation of the RFI with ultraviolet light markedly increases the rate of hydrolysis by A(3). B appears to form 3'-phosphoryl termini with fd DNA, but its characterization is highly preliminary due to its instability.

Chemical Precipitation↗