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Datasheet for Histone H3.1 Human, Recombinant (M2503; Lot 0041312)

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Datasheet for Histone H3.1 Human, Recombinant (M2503; Lot 0041312) Source: An E. coli strain that carries a plasmid Note: The protein concentration (1 mg/ml, 65 µM) weight on 10–20% Tris-Glycine SDS-PAGE gel is encoding the cloned human histone H3.1 gene, is calculated using the molar extinction coefficient ~15 kDa. Histone H3.1 HIST1H3A or H3FA. (Genbank accession number: for Histone H3.1 (4080) and its absorbance at Mass Spectrometry: The mass of purified AF531274) 280 nm (3,4). 1.0 A units = 3.8 mg/ml Human, Recombinant 280 Histone H3.1 Human, Recombinant is 15273.2 Da Supplied in: 20 mM Sodium Phosphate (pH 7.0), Synonym: Histone H3/a. as determined by ESI-TOF MS (Electrospray 1-800-632-7799 [email protected] 300 mM NaCl, 1 mM EDTA and 1 mM DTT. Ionization-Time of Flight Mass Spectrometry). The www.neb.com Quality Control Assays: average mass calculated from primary sequence M2503S 004131215121 SDS-PAGE: 0.5, 1.0, 2.0, 5.0, 10.0 µg of Histone is 15272.89 Da. This confirms the protein identity H3.1 Human, Recombinant were loaded on a as well as the absence of any modifications of the B r 1 2 3 4 5 6 7 M2503S kDa 10–20% Tris-Glycine SDS-PAGE gel and stained histone. 250 with Coomassie Blue. The calculated molecular 100 µg 1.0 mg/ml Lot: 0041312 N-terminal Protein Sequencing: Protein identity 150 weight is 15272.89 Da. Its apparent molecular 100 was confirmed using Edman Degradation to RECOMBINANT Store at –20°C Exp: 12/15 15273.2 80 100 sequence the intact protein. Description: Histone H3 combines with Histone 60 90 50 Enzyme Modification: H4 to form the H3/H4 tetramer. Two H2A/H2B 80 40 1. G9a Methyltransferase: After incubation of a heterodimers interact with an H3/H4 tetramer to 70 form the histone octamer (1,2). It is also modified 30 25 µl reaction for 10 minutes at 37°C, 1 unit 60 by various enzymes and can act as a substrate for 25 of G9a methyltransferase (NEB #M0235) 50 transfers 0.6 pmols of methyl group to Histone them. These modifications have been shown to be 20 % Intensity 40 H3.1 Human, Recombinant. important in gene regulation. 15 30 2. SET7 Methyltransferase: After incubation of a Histone H3.1, an H3 variant that has thus far only SDS-PAGE analysis of Histone H3.1 Human, Recombinant. 20 25 µl reaction for 10 minutes at 37°C, 1 unit been found in mammals, is replication dependent of SET7 methyltransferase (NEB #M0233) and is associated with gene activation and gene Lane 1 & 7: NEB Protein Ladder (NEB #P7703), Lane 2 thru 10 6: 0.5–10.0 µg Histone H3.1 Human, Recombinant. (Please transfers 1 pmol of methyl group to Histone silencing (3). see Quality Control section for more information) 10000 12000 14000 16000 18000 20000 H3.1 Human, Recombinant. Mass, Da ESI-TOF Analysis of Histone H3.1 Human, Recombinant. (See other side) CERTIFICATE OF ANALYSIS Source: An E. coli strain that carries a plasmid Note: The protein concentration (1 mg/ml, 65 µM) weight on 10–20% Tris-Glycine SDS-PAGE gel is encoding the cloned human histone H3.1 gene, is calculated using the molar extinction coefficient ~15 kDa. Histone H3.1 HIST1H3A or H3FA. (Genbank accession number: for Histone H3.1 (4080) and its absorbance at Mass Spectrometry: The mass of purified AF531274) 280 nm (3,4). 1.0 A units = 3.8 mg/ml Human, Recombinant 280 Histone H3.1 Human, Recombinant is 15273.2 Da Supplied in: 20 mM Sodium Phosphate (pH 7.0), Synonym: Histone H3/a. as determined by ESI-TOF MS (Electrospray 1-800-632-7799 [email protected] 300 mM NaCl, 1 mM EDTA and 1 mM DTT. Ionization-Time of Flight Mass Spectrometry). The www.neb.com Quality Control Assays: average mass calculated from primary sequence M2503S 004131215121 SDS-PAGE: 0.5, 1.0, 2.0, 5.0, 10.0 µg of Histone is 15272.89 Da. This confirms the protein identity H3.1 Human, Recombinant were loaded on a as well as the absence of any modifications of the B r 1 2 3 4 5 6 7 M2503S kDa 10–20% Tris-Glycine SDS-PAGE gel and stained histone. 250 with Coomassie Blue. The calculated molecular 100 µg 1.0 mg/ml Lot: 0041312 N-terminal Protein Sequencing: Protein identity 150 weight is 15272.89 Da. Its apparent molecular RECOMBINANT Store at –20°C Exp: 12/15 100 was confirmed using Edman Degradation to 15273.2 80 100 sequence the intact protein. Description: Histone H3 combines with Histone 60 90 50 Enzyme Modification: H4 to form the H3/H4 tetramer. Two H2A/H2B 80 40 1. G9a Methyltransferase: After incubation of a heterodimers interact with an H3/H4 tetramer to 70 form the histone octamer (1,2). It is also modified 30 25 µl reaction for 10 minutes at 37°C, 1 unit 60 by various enzymes and can act as a substrate for 25 of G9a methyltransferase (NEB #M0235) 50 transfers 0.6 pmols of methyl group to Histone them. These modifications have been shown to be % Intensity 20 important in gene regulation. 40 H3.1 Human, Recombinant. 15 30 2. SET7 Methyltransferase: After incubation of a Histone H3.1, an H3 variant that has thus far only 20 25 µl reaction for 10 minutes at 37°C, 1 unit been found in mammals, is replication dependent SDS-PAGE analysis of Histone H3.1 Human, Recombinant. of SET7 methyltransferase (NEB #M0233) and is associated with gene activation and gene Lane 1 & 7: NEB Protein Ladder (NEB #P7703), Lane 2 thru 10 6: 0.5–10.0 µg Histone H3.1 Human, Recombinant. (Please transfers 1 pmol of methyl group to Histone silencing (3). see Quality Control section for more information) 10000 12000 14000 16000 18000 20000 H3.1 Human, Recombinant. Mass, Da ESI-TOF Analysis of Histone H3.1 Human, Recombinant. (See other side) CERTIFICATE OF ANALYSIS Protease Assay: After incubation of 10 µg of Histone Protein Sequence: ARTKQTARKSTGGKAPRK H3.1 Human, Recombinant with a standard mixture QLATKAARKSAPATGGVKKPHRYRPGTVALREI of proteins for 2 hours at 37°C, no proteolytic RRYQKSTELLIRKLPFQRLVREIAQDFKTDLRFQ activity could be detected by SDS-PAGE. SSAVMALQEACEAYLVGLFEDTNLCAIHAKRVT Exonuclease Assay: Incubation of a 50 µl IMPKDIQLARRIRGERA (Genbank accession reaction containing 10 µg of Histone H3.1 Human, number: AAN10051) Recombinant with 1 µg of a mixture of single and Usage Note: When running SDS-PAGE gels, double-stranded [3H] E. coli DNA (200,000 cpm/ Histone H3.1 can self-oligomerize if freshly µg) for 4 hours at 37°C released < 0.1% of the total prepared DTT (dithiothreitol) is not used when radioactivity. preparing the sample for gel electrophoresis Endonuclease Assay: Incubation of a 50 µl References: reaction containing 10 µg of Histone H3.1 Human, 1. Kornberg, R.D. (1977) Annu. Rev. Biochem. Recombinant with 1 µg of φX174 RF I (suprecoiled) 46, 931–954. plasmid DNA for 4 hours at 37°C resulted in 2. van Holde, K.E. (1989) Chromatin, 1–497. < 5.0% conversion to RF II form (nicked circle) as determined by agarose gel electrophoresis. 3. Hake, S.B. et al. (2006) J. Biol. Chem., 281, 559–568. 4. Gill, S.C. and von Hippel, P.H. (1989) Anal. Biochem. 182, 319–326. 5. Pace, C.N. et al. (1995) Protein Science, 4, 2411–2423. Page 2 (M2503) Protease Assay: After incubation of 10 µg of Histone Protein Sequence: ARTKQTARKSTGGKAPRK H3.1 Human, Recombinant with a standard mixture QLATKAARKSAPATGGVKKPHRYRPGTVALREI of proteins for 2 hours at 37°C, no proteolytic RRYQKSTELLIRKLPFQRLVREIAQDFKTDLRFQ activity could be detected by SDS-PAGE. SSAVMALQEACEAYLVGLFEDTNLCAIHAKRVT Exonuclease Assay: Incubation of a 50 µl IMPKDIQLARRIRGERA (Genbank accession reaction containing 10 µg of Histone H3.1 Human, number: AAN10051) Recombinant with 1 µg of a mixture of single and Usage Note: When running SDS-PAGE gels, double-stranded [3H] E. coli DNA (200,000 cpm/ Histone H3.1 can self-oligomerize if freshly µg) for 4 hours at 37°C released < 0.1% of the total prepared DTT (dithiothreitol) is not used when radioactivity. preparing the sample for gel electrophoresis Endonuclease Assay: Incubation of a 50 µl References: reaction containing 10 µg of Histone H3.1 Human, 1. Kornberg, R.D. (1977) Annu. Rev. Biochem. Recombinant with 1 µg of φX174 RF I (suprecoiled) 46, 931–954. plasmid DNA for 4 hours at 37°C resulted in 2. van Holde, K.E. (1989) Chromatin, 1–497. < 5.0% conversion to RF II form (nicked circle) as determined by agarose gel electrophoresis. 3. Hake, S.B. et al. (2006) J. Biol. Chem., 281, 559–568. 4. Gill, S.C. and von Hippel, P.H. (1989) Anal. Biochem. 182, 319–326. 5. Pace, C.N. et al. (1995) Protein Science, 4, 2411–2423. Page 2 (M2503).
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