Photochemical evidence for conformation changes in DNA during germination of bacterial spores.

Stafford, Robert; Donnellan, J. Edward

doi:10.1073/pnas.59.3.822

Cited by 66 publications

(32 citation statements)

References 10 publications

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“…It has been suggested that small regions of DNA are in an A-like conformation in vivo thus facilitating interaction with specific proteins, but this suggestion has never been substantiated (Reynolds et ai., 1985;Rhodes and Klug, 1986). However, it was proposed almost 25 years ago that DNA in dormant spores of Baciiius species is entirely in an A-like conformation (Stafford and Donnellan, 1968). While one of the assumptions on which this proposal was based turned out to be incorrect, I believe the proposal as originally made was correct.…”

Section: Dna Conformation In Vitro and In Vivomentioning

confidence: 80%

“…Normally, a/p-type SASP are degraded owing to the action of a SASP-specific protease in the first minutes of spore germination (Setlow, 1988a), at which time plasmid supercoiling and the u.v. photochemistry of DNA return to that of growing cells (Table 1) (Nicholson and Setlow, 1990;Stafford and Donnellan, 1968). There is, however, a transient change in the u.v.…”

Section: Small Acid-soluble Proteins Involved In A-dna Formation In Bmentioning

confidence: 99%

“…There is, however, a transient change in the u.v. photochemistry of DNA in the first minutes of spore germination, as there is a brief period when no photoproducts are produced (Stafford and Donnellan, 1968). This time period may include the actual conformational°l f this increased level of negative supercoiling is largely due to a B -» A conformational change in the DNA, then there may actually be no change in the superhelical tension of DNA in spores relative to that in cells (Driica, 1992).…”

Section: Small Acid-soluble Proteins Involved In A-dna Formation In Bmentioning

confidence: 99%

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DNA in dormant spores of Bacillus species is in an A‐like conformation

Setlow

1992

Molecular Microbiology

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SummaryThe DNA in dormant spores of Bacillus species is associated with a/|3-type smaii, acid-soiuble proteins (SASP), which are double-stranded DNA-binding proteins whose amino acid sequence has been highly conserved in evolution. In vitro these proteins bind most strongly to DNA which readily adopts an A-iike conformation, as binding of a/|3-type SASP causes DNA to assume an A-like conformation. As predicted by this conformational change in DNA, binding of a/ptype SASP to relaxed but covalently closed plasmid DNA results in the introduction of a iarge number of negative supercoils. Associated with the conformational change in DNA brought about by oc/^-type SASP binding is a change in its photochemistry such that ultraviolet irradiation does not generate pyrimidine dimers, but rather a thyminyl-thymine adduct termed spore photoproduct (SP). The latter two properties of DNA complexed with o(/|3-type SASP in vitro are similar to those of DNA in dormant spores of Baciiius species in which: (i) plasmid DNA has a much higher number of negative supercoils than plasmid in growing cells; and (ii) ultraviolet irradiation produces SP and no pyrimidine dimers, while only pyrimidine dimers are formed in growing cells. During sporulation these changes in the properties of spore DNA take place in parallel with synthesis of a/ptype SASP, and the magnitude of the changes is greatly reduced in mutants that make low amounts of these proteins. A straightforward interpretation of these data is that DNA in dormant spores of Bacillus species is in an A-like conformation. DNA in an A-like conformation is also present in growing cells with high a/|3-type SASP levels. However, the presence of this A-like DNA is detrimental to cell growth.

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Section: Dna Conformation In Vitro and In Vivomentioning

confidence: 80%

Section: Small Acid-soluble Proteins Involved In A-dna Formation In Bmentioning

confidence: 99%

Section: Small Acid-soluble Proteins Involved In A-dna Formation In Bmentioning

confidence: 99%

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DNA in dormant spores of Bacillus species is in an A‐like conformation

Setlow

1992

Molecular Microbiology

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“…Since UV irradiation of purified DNA under conditions of low humidky also forms a significant amount of spore photoproduct [10], the altered DNA photochemistry in spores has been attributed to an alteration in DNA conformation: when the DNA undergoes a change from B-form (at high relative humidity) to A-form (at lower humidity) its photoproduct distribution switches from one consisting mostly of thymine dimers to one dominated by spore photoproduct. This leads to the obvious infemnc~ that ~,type SASP may provide a UV-protective effect to spores by causing a B ---> A conformation change upon binding to DNA [11,12], In vitro studies by Mohr et al [13] have borne out this supposition: both circular dichroism (CD) and Fourier-transform infrared (FTIR) spectroscopy demonstrate the proposed conformation change upon ~/fl-type SASP binding. -10 -5 1 5 10 15 20 25 30 (gl-6) 35 40 45 50 55 60 I l I I I I I I [ I I 1 I I We have now embarked on structural studies aimed at elucidating the mechanism of action of this novel family of DNA binding proteins.…”

Section: Introductionmentioning

confidence: 99%

Synthesis and characterization of a 29‐amino acid residue DNA‐binding peptide derived from α/β‐type small, acid‐soluble spore proteins (SASP) of bacteria

et al. 1992

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A 29-amino acid residue peptide (SASP-peptide) derived from the sequence of the putative DNA-eontacting portion at the carboxyi terminus of an g/fl-type small, acid-soluble spore protein (SASP) of Bacillus subtllfs has been synthesized by automated solid-plms¢ methods and testcgt for its ability to interact with DNA. Circular dichroism (CD) spectroscopy reveals an interaction between this SASP-peptid¢ and DNA, both by an increase in a.helix content of the peptide (which alone has a mostly random conformation) and by enhancement of the 27~-nm CD band of the DNA. In contrast to results with intact a/a-type SASP, however, the peptide does not induce a B ~ A co,formational transition in the DNA. The SASP.peptide also binds to poly(dG), poly(dC) and protects this polynucleotide against DNas¢ I digestion and UV light.induced cytosine dimer formation, parallel to findings made previously with native g/fl.type SASP. The results confirm the hypothesis that the carboxyl-terminal region of the g/p.type SASF directly contacts DNA and possesses some, but not all. of the functional characteristics of the intact molecule.

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“…Resistance of bacterial spores to ultraviolet ray (UV) increases temporarily in the course of germination (1)(2)(3)(4). STAFFORD and DONNELLAN (3) showed that this increase of UV-resistance in germinating spores was correlated with a decrease in the production of thymine-containing photoproducts.…”

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confidence: 99%

Uv-Resistance of Transforming Dna in Germinating Spores of Bacillus Subtilis

Irie

Yano

Kembo

1970

J. Gen. Appl. Microbiol.

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Resistance of bacterial spores to ultraviolet ray (UV) increases temporarily in the course of germination (1-4). STAFFORD and DONNELLAN (3) showed that this increase of UV-resistance in germinating spores was correlated with a decrease in the production of thymine-containing photoproducts. The experiments described below were carried out to see if this increase in UV-resistance could be explained in terms of the resistance of DNA in the spores as measured by transforming ability. Spores of Bacillus subtilis Marburg 160 try-arg-were prepared as described by TANOOKA (5). To facilitate the extraction of DNA from spores (6), it was necessary to sensitize the spores to lysozyme (7) prior to germination and UV-irradiation.The sensitization to lysozyme was performed by treating the dormant spores (about 109 spores/ml) at 37° for 90 min with 10% thioglycolic acid in the presence of 7.2 M urea and 0.5 mM dithiothreitol (8). The spores thus treated were centrifuged, washed twice with 0.1 M phosphate buffer (pH 7.8), and suspended in distilled water. At this stage, the spores were highly dormant and required heat-shock for germination.After being heat-shocked at 70° for 2 hr, the spores (about 109 spores/ ml) were germinated at 37° in 10 mM Tris buffer (pH 7.0) containing 10 mM L-alanine. As shown in Fig. 1, these spores exhibited an unusually long lag period (30 min or more at this spore concentration) before commencement of germination as measured by optical density.This lag period tended to increase with spore concentration.Despite these abnormal behaviors caused by the thioglycolic acid treatment, germination of these spores was considered to be physiological in nature because (1) they were fully viable and (2) no appreciable change in optical density occurred in the absence of alanine.For the examination of UV-resistance of DNA in spores, an aliquot (5 ml) removed from the germinating spore suspension was chilled quickly to below 10°, transferred to an aluminium dish (76 mm in diameter) and sub-181

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Photochemical evidence for conformation changes in DNA during germination of bacterial spores.

Cited by 66 publications

References 10 publications

DNA in dormant spores of Bacillus species is in an A‐like conformation

DNA in dormant spores of Bacillus species is in an A‐like conformation

Synthesis and characterization of a 29‐amino acid residue DNA‐binding peptide derived from α/β‐type small, acid‐soluble spore proteins (SASP) of bacteria

Uv-Resistance of Transforming Dna in Germinating Spores of Bacillus Subtilis

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