Chromatin acidic proteins with high affinity for nucleohistone and DNA

Patel, Gordhan L.

doi:10.1016/0024-3205(72)90268-8

Cited by 26 publications

(8 citation statements)

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“…The majority of proteins in this fraction migrate as two major bands. Their molecular weight has been estimated to be 16,300 and 19,600 (14). The relative affinities of the various fractions of nuclear acidic protein for rat-liver DNA are presented in Table 1.…”

Section: Resultsmentioning

confidence: 99%

“…Three acidic protein fractions were extracted from purified nuclei on the basis of their solubility properties and ability to reconstitute with the deoxynucleoprotein chromatin. Details of the extraction procedure have been reported elsewhere (14). Briefly, nuclei were extracted successively with 0.14 M NaCl-50 mM Trisc HCl (pH 7.5) to obtain the proteins of the nuclear sap (NSP), and with 2 M NaCl-5 M urea-10 mM Tris * HCl (pH 8) to obtain a soluble chromatin fraction, which was dialyzed overnight against 13 volumes of distilled water to lower the NaCl concentration to 0.14 M. This resulted in precipitation of the reconstituted chromatin, leaving in the soluble phase a fraction of chromatin acidic proteins (CAP-I) irreversibly dissociated by the high-ionic-strength medium (3).…”

Section: Methodsmentioning

confidence: 99%

“…Our knowledge of even the elementary properties of these chromatin acidic proteins, however, is meager due to the technical difficulties encountered in their isolation, fractionation, and characterization (2). Chromatin a9eidic proteins, isolated by various methods from many cell types, are heterogeneous and tissue and species specific (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17) , and confer tissue and species specificity to the chromatin template in transcription (8,(18)(19)(20). The latter observations strongly indicate a gene regulatory role for at least some of the nonhistone acidic proteins.…”

Section: Introductionmentioning

confidence: 99%

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Some Binding Parameters of Chromatin Acidic Proteins with High Affinity for Deoxyribonucleic Acid

Patel¹,

Thomas²

1973

Proc. Natl. Acad. Sci. U.S.A.

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A fraction of chromatin acidic protein from rat liver, predominantly comprised of low-molecular-weight species, showed highest in vitro affinity for DNA when compared with other fractions of nuclear acidic protein. Maximum binding of this protein fraction was observed at physiological ionic strength of 0.14 M NaCi and DNA/protein ratio > 1. Comparison of their binding to homologous and heterologous DNAs showed partial species specificity. All molecular species of this protein fraction appear to bind to DNA.A prerequisite feature of proteins involved in the structure and function of the genome is the capacity to interact specifically with deoxyribonucleic acid. In eukaryotes, histones, a class of basic proteins, which have been recognized as DNA-associated components of the chromatin, have been investigated intensely (ref. 1 for recent review); and yet, their exact nuclear function has not been clearly defined. In addition to histones, the chromatin also contains a variable, but significant, amount of acidic proteins. Our knowledge of even the elementary properties of these chromatin acidic proteins, however, is meager due to the technical difficulties encountered in their isolation, fractionation, and characterization (2). Chromatin a9eidic proteins, isolated by various methods from many cell types, are heterogeneous and tissue and species specific (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17) , and confer tissue and species specificity to the chromatin template in transcription (8,(18)(19)(20). The latter observations strongly indicate a gene regulatory role for at least some of the nonhistone acidic proteins. Some recent reports have shown in vitro binding of some chromatin acidic proteins to DNA in a species-specific manner (11,21). These observations are analogous to those in prokaryotes, where gene regulatory proteins interact specifically with DNA (22,23). A study of nuclear proteins, in particular the nonhistone acidic proteins of chromatin with high affinity for DNA, is, therefore, of interest in connection with their possible role in chromatin structure and/or in regulation of replication and transcription.Recently the isolation from rat-liver chromatin of a class of acidic proteins with high affinity for nucleohistone and DNA and predominantly comprised of low-molecular-weight species was reported from this laboratory (14). These proteins are distinct from other fractions of nuclear acidic proteins and from chromatin acidic proteins that bind DNA reported by Acidic Proteins with High others. In the present study we report on some of the in vitro DNA binding parameters of these acidic proteins. MATERIALS AND METHODSIsolation of Nuclei. Male Sprague-Dawley rats (200-250 g) were kept on Purina chow freely until 24 hr before they were killed by decapitation. Livers were removed immediately, washed with ice-cold 0.25 M sucrose-3 mM CaCl2, and processed to purify nuclei by the method of Pogo et al. (24), with additional filtration of tissue homogenates through Miracloth and substitution of Mg...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Some Binding Parameters of Chromatin Acidic Proteins with High Affinity for Deoxyribonucleic Acid

Patel¹,

Thomas²

1973

Proc. Natl. Acad. Sci. U.S.A.

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“…In our laboratory, we have isolated a distinct subclass of the NHCP, designated APNH (acidic proteins with affinity for nucleohistone) (18), and have studied its in vitro DNA binding properties. We have demonstrated that APNH binds to DNA optimally at physiological conditions (9)(10)(11).…”

Section: Helix-coil Transition Of Synthetic Poly~d(a-t)d(a-t)j At 25°mentioning

confidence: 99%

“…This subfraction contains about 10% of the total NHCP and precipitates with the deoxynucleohistone when chromatin solution in buffered 2 M NaCl and 5 M urea is dialyzed against 13 volumes of distilled water. It is separated from histones and DNA by chromatography on hydroxylapatite (18 (20) and by the Lowry et al method (21). Polyacrylamide gel electrophoresis in sodium dodecyl sulfate of proteins was as described by MacGillivray et al (22) (19).…”

Section: That Unwinds Synthetic Poly[d(a-t)-d(a-t)] At Room Temperatumentioning

confidence: 99%

DNA unwinding component of the nonhistone chromatin proteins.

Thomas¹,

Patel²

1976

Proc. Natl. Acad. Sci. U.S.A.

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A subclass of nonhistone chromatin proteins from rat liver, previously shown to exhibit high affinity for DNA, has been fractionated by single-stranded DNA-agarose affinity chromatography. The protein fraction that bound to DNAagarose in 0.19 M NaCl-buffer and was eluted with 2 M NaClbuffer is enriched for a protein component of approximately 20,000 daltons and exhibits preferential binding to denatured DNA. This nonhistone protein fraction specific for single strands binds to DNA in a non-species-specific manner, and causes helix-coil transition of synthetic poly~d(A-T)d(A-T)J at 25°, as indicated by the increase in absorbance of ultraviolet light at 260 nm. The observed hyperchromicity does not result from any nuclease activity in the protein fraction, because addition of Mg+2 results in partial hypochromic shift, and the protein/DNA complex is retained by nitrocellulose filters. Protein/DNA interactions are clearly important in the structure and function of the eukaryotic chromatin. Interactions of histones with DNA have been extensively analyzed and, in recent years, have been shown to give rise to the subunit structure of the chromatin (1-5). The role of the nonhistone chromatin proteins (NHCP) in the chromatin is also a subject of much current interest. However, the NHCP are a heterogeneous class of proteins that have long resisted fractionation and characterization. Recently, the DNA binding properties of the NHCP have been exploited to fractionate and characterize them, and to gain insight into their possible biological roles (6-17).In our laboratory, we have isolated a distinct subclass of the NHCP, designated APNH (acidic proteins with affinity for nucleohistone) (18), and have studied its in vitro DNA binding properties. We have demonstrated that APNH binds to DNA optimally at physiological conditions (9-11). Equilibrium and kinetic competition experiments indicated that this protein fraction interacts preferentially with A-T rich and/or singlestranded DNA, which suggested that APNH might affect the helix-coil transition of DNA (11)

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Nagl

1973

Fortschritte Der Botanik

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Chromatin acidic proteins with high affinity for nucleohistone and DNA

Cited by 26 publications

References 10 publications

Some Binding Parameters of Chromatin Acidic Proteins with High Affinity for Deoxyribonucleic Acid

Some Binding Parameters of Chromatin Acidic Proteins with High Affinity for Deoxyribonucleic Acid

DNA unwinding component of the nonhistone chromatin proteins.

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