NORMAL GENOMIC

• Saitoh F, Kawamura S, Yamasaki N, Tanaka I, Kimura M
• Arginine-55 in the beta-arm is essential for the activity of DNA-binding protein HU from Bacillus stearothermophilus.
• Biosci Biotechnol Biochem. 1999; 63: 2232-5
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• DNA-binding protein HU (BstHU) from Bacillus stearothermophilus is a homodimeric protein which binds to DNA in a sequence-nonspecific manner. In order to identify the Arg residues essential for DNA binding, four Arg residues (Arg-53, Arg-55, Arg-58, and Arg-61) within the beta-arm structure were replaced either by Gln, Lys, or Glu residues, and the resulting mutants were characterized with respect to their DNA-binding activity by a filter-binding analysis and surface plasmon resonance analysis. The results indicate that three Arg residues (Arg-55, Arg-58, and Arg-61) play a crucial role in DNA binding as positively charged recognition groups in the order of Arg-55 > Arg-58 > Arg-61 and that these are required to decrease the dissociation rate constant for BstHU-DNA interaction. In contrast, the Arg-53 residue was found to make no contribution to the binding activity of BstHU.

• Vis H, Dobson CM, Robinson CV
• Selective association of protein molecules followed by mass spectrometry.
• Protein Sci. 1999; 8: 1368-70
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• Nanoflow electrospray mass spectrometry was used to monitor the formation of protein heterodimers of HU proteins from Bacillus stearothermophilus and Bacillus subtilis. This has enabled us to analyze both thermodynamic and kinetic features associated with the dissociation of homodimeric HU proteins. The results obtained correlate well with the kinetics of the protein dissociation process and the free energy difference between homo- and heterodimeric species anticipated from other studies. We suggest that this approach will have general applicability in studying protein association and dissociation under near-equilibrium conditions and will be relevant to a wide range of biological systems.

• Tominaga T, Nakagawa A, Tanaka I, Kawamura S, Kimura M
• High-resolution crystals of the HU mutant K38N from Bacillus stearothermophilus.
• J Struct Biol. 1999; 125: 86-9
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• The DNA-binding protein HU is ubiquitous in the prokaryotic cell. It is a major protein component of isolated nucleoids and is believed to control the tertiary structure of prokaryotic DNA. The Bacillus stearothermophilus HU (BstHU) mutants obtained by mutagenesis have been investigated. Crystallization experiments of BstHU-K38N (Lys38 is substituted with Asn) resulted in two forms of crystals suitable for high-resolution x-ray analysis. The first form belongs to the monoclinic space group C2 with unit-cell dimensions of a = 90.1 A, b = 43.5 A, c = 63.7 A, and beta = 135.1 degrees, and it diffracts x rays to 1.5-A resolution. The second form belongs to the tetragonal space group I41 with a = b = 62.6 A and c = 43.3 A, and it diffracts up to 1.8-A resolution.

• Laine B, Chartier F, Culard F, Belaiche D, Sautiere P
• The DNA-binding protein II from Zymomonas mobilis. Complete amino acid sequence and interaction with DNA.
• Biochimie. 1998; 80: 109-16
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• The primary structure of the DNA-binding protein II from Zymomonas mobilis has been determined from data provided by automated Edman degradation of the intact protein and of peptides derived from cleavage at aspartic acid and arginine residues. When compared with the homologous protein isolated from other bacteria, the DNA-binding protein II from Z mobilis shows many substitutions. Several non-conservative substitutions at positions usually highly conserved in this type of protein probably account for the weaker DNA-binding activity of this protein compared to that of the E coli protein.

• Rice PA
• Making DNA do a U-turn: IHF and related proteins.
• Curr Opin Struct Biol. 1997; 7: 86-93
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• IHF and HU belong to a family of proteins that introduce sharp bends into DNA and act as accessory factors in a variety of cellular processes in prokaryotes. In addition to the crystal structure of IHF bound to DNA, the past year has seen a number of advances in the understanding of the interactions of these proteins with DNA in solution.

• Ellenberger T, Landy A
• A good turn for DNA: the structure of integration host factor bound to DNA.
• Structure. 1997; 5: 153-7
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• The crystal structure of integration host factor (IHF) complexed with DNA shows how a small heterodimeric protein can induce a big bend in DNA. IHF exerts leverage in the minor groove and wraps DNA around the body of the protein, providing another example of sequence-specific recognition of the minor groove.

• Metallo SJ, Schepartz A
• Certain bZIP peptides bind DNA sequentially as monomers and dimerize on the DNA.
• Nat Struct Biol. 1997; 4: 115-7

• Ogata K, Nishimura Y
• [Mechanism of DNA recognition by a protooncogene product, Myb]
• Seikagaku. 1996; 68: 111-28

• Boelens R, Vis H, Vorgias CE, Wilson KS, Kaptein R
• Structure and dynamics of the DNA binding protein HU from Bacillus stearothermophilus by NMR spectroscopy.
• Biopolymers. 1996; 40: 553-9
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• The DNA-binding protein HU from Bacillus stearothermophilus (HUBst) is a dimer with a molecular weight of 195 kDa that is capable of bending DNA. An x-ray structure has been determined previously [Tanaka et al. 1984) Nature, vol. 310, pp. 376-381], but no structure could be established for a large part of the supposed DNA-binding beta-arms. Distance geometry and restrained molecular dynamics using nmr restraints were used to generate a set of 25 structures. These structures display a backbone rms deviation (RMSD) of 0.36 A for the well-defined region (residues 2-54 and 75-90). The structure of the core is very similar to that observed in the x-ray structure, with a pairwise RMSD of 1.06 A. The structure of the beta-hairpin arm contains a double flip-over at the prolines in the two strands of the beta-arm. Heteronuclear 15N relaxation measurements indicate that the beta-arm and the tip of the beta-arm is flexible. This explains the disorder observed in the solution and x-ray structures of the beta-arm with respect to the core of the protein. Overlayed onto itself the beta-arm is better defined, with a backbone RMSD of 1.0 A calculated for residues 54-59 and 69-74. The tip of the arm adopts a well-defined 4 : 6 beta-hairpin conformation. Changes in amide 15N and 1H chemical shifts upon titrating DNA are most pronounced for the residues in the beta-hairpin arm and for the residues in the second half of the third alpha-helix. Heteronuclear 15N relaxation data for free and complexed HUBst show that that the arms become structured upon DNA binding. Together with chemically induced nuclear polarization measurements on a mutant HUBst (M69Y; V76Y) this shows that the beta-hairpin arm is involved in direct DNA interaction.

• Grayling RA, Sandman K, Reeve JN
• DNA stability and DNA binding proteins.
• Adv Protein Chem. 1996; 48: 437-67

• Lunsford RD, Nguyen N, London J
• DNA-binding activities in Streptococcus gordonii: identification of a receptor-nickase and a histonelike protein.
• Curr Microbiol. 1996; 32: 95-100
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• Extraction of Streptococcus gordonii cells with the mild chaotropic agent, LiCl, drastically decreased DNA transforming ability, had little effect on viability, and released both DNA nicking and binding activities. Both activities were Mg2+ and Ca2+ independent and were not competence specific. Southwestern blot analysis of the extract identified putative surface proteins of 56 kDa and 68 kDa in strain Challis and Wicky, respectively. Extracts also contained a 10-kDa DNA-binding protein, designated HSgo, that belongs to the eubacterial histonelike class of proteins.

• Parkinson G et al.
• Aromatic hydrogen bond in sequence-specific protein DNA recognition.
• Nat Struct Biol. 1996; 3: 837-41

• Hayat MA, Mancarella DA
• Nucleoid proteins.
• Micron. 1995; 26: 461-80
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• This article examines the published evidence in support of the classification of organisms into three groups (Bacteria, Archae, and Eukarya) instead of two groups (prokaryotes and eukaryotes) and summarizes the comparative biochemistry of each of the known histone-like, nucleoid DNA-binding proteins. The molecular structures and amino acid sequences of Archae are more similar to those of Eukarya than of Bacteria, with a few exceptions. Cytochemical methodology employed for localizing these proteins in archaeal and bacterial cells has also been reviewed. It is becoming increasingly apparent that these proteins participate both in the organization of DNA and in the control of gene expression. Evidence obtained from biochemical properties, structural and functional differences, and the ultrastructural location of these proteins, as well as from gene mutations clearly justifies the division of prokaryotes into bacterial and archaeal groups. Indeed, chromosomes, whether they be nuclear, prokaryotic, or organellar, are invariably complexed with abundant, small, basic proteins that bind to DNA with low sequence specificity. These proteins include the histones, histone-like proteins, and nonhistone high mobility group (HMG) proteins.

• von Hippel PH
• Protein-DNA recognition: new perspectives and underlying themes.
• Science. 1994; 263: 769-70

• Baumann H, Knapp S, Lundback T, Ladenstein R, Hard T
• Solution structure and DNA-binding properties of a thermostable protein from the archaeon Sulfolobus solfataricus.
• Nat Struct Biol. 1994; 1: 808-19
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• The archaeon Sulfolobus solfataricus expresses large amounts of a small basic protein, Sso7d, which was previously identified as a DNA-binding protein possibly involved in compaction of DNA. We have determined the solution structure of Sso7d. The protein consists of a triple-stranded anti-parallel beta-sheet onto which an orthogonal double-stranded beta-sheet is packed. This topology is very similar to that found in eukaryotic Src homology-3 (SH3) domains. Sso7d binds strongly (Kd < 10 microM) to double-stranded DNA and protects it from thermal denaturation. In addition, we note that epsilon-mono-methylation of lysine side chains of Sso7d is governed by cell growth temperatures, suggesting that methylation is related to the heat-shock response.

• Imbert M, Laine B, Helbecque N, Mornon JP, Henichart JP, Sautiere P
• Conformational study of the chromosomal protein MC1 from the archaebacterium Methanosarcina barkeri.
• Biochim Biophys Acta. 1990; 1038: 346-54
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• Methanogen chromosomal protein MC1 is a polypeptide of 93 amino acid residues (Mr 10,757) which represents the major protein associated with the DNA of the archaebacterium Methanosarcina barkeri and can protect DNA against thermal denaturation. The conformation of protein MC1 has been investigated by means of predictive methods, infrared spectroscopy, circular dichroism and tryptophan fluorescence studies. Protein MC1 has a low amount of alpha-helix but contains antiparallel beta-sheet strands. The larger hydrophobic cluster which contains tryptophan at position 61 appears buried in the protein. Addition of salts induces the unfolding of the protein and makes the tryptophan indole ring more rigid. With respect to its primary structure and its conformation, protein MC1 appears radically different from the chromosomal DNA-binding protein II (also called HU-type protein) in eubacteria.

• Bruckner RC, Cox MM
• The histone-like H protein of Escherichia coli is ribosomal protein S3.
• Nucleic Acids Res. 1989; 17: 3145-61
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• We report the purification of four proteins from Escherichia coli that stimulate or inhibit inter- and/or intramolecular recombination promoted by the yeast plasmid-encoded FLP protein. The proteins are identified as the ribosomal proteins S3 (27 kDa), L2 (26 kDa), S4 (24 kDa), and S5 (16 kDa), on the basis of N-terminal sequence analysis. The S3 protein is found to be identical to H protein, an E. coli histone-like protein that is related to histone H2A immunologically and by virtue of amino acid content. The H protein/S3 identity is based on co-migration on polyacrylamide gels, heat stability, amino acid analysis, and effects on FLP-promoted recombination. These results are relevant to current studies on the structure of the E. coli nucleoid. Since the H protein has previously been found associated with the E. coli nucleoid, the results indicate that either (a) some ribosomal proteins serve a dual function in E. coli, or, more likely, (b) ribosomal proteins can and are being mis-identified as nucleoid constituents.

• Reddy TR, Suryanarayana T
• Archaebacterial histone-like proteins. Purification and characterization of helix stabilizing DNA binding proteins from the acidothermophile Sulfolobus acidocaldarius.
• J Biol Chem. 1989; 264: 17298-308
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• Four DNA binding histone-like proteins have been purified from the nucleoid of the acidothermophilic archaebacterium Sulfolobus acidocaldarius to homogeneity employing DNA-cellulose chromatography and carboxymethylcellulose chromatography. The molecular weights of these proteins are in the range 8,000-12,500. Immunoblotting results suggest that a few antigenic determinants are common among these proteins which could not be detected by immunodiffusion. Spectroscopic properties of the proteins have been studied. The amino acid compositions of these proteins show both similarities and differences with histones and prokaryotic histone-like proteins. All of the four proteins bind native and denatured DNAs and single stranded RNA with differing affinities. Three of the proteins, denoted by HSNP (helix stabilizing nucleoid protein)-A, HSNP-C, and HSNP-C', show physiologically significant, strong, and synergistic effects in stabilizing duplex DNA against thermal denaturation with Tm increases in the range of 15-30 +/- degrees C.

• Pettijohn DE
• Histone-like proteins and bacterial chromosome structure.
• J Biol Chem. 1988; 263: 12793-6

• Falconi M, Gualtieri MT, La Teana A, Losso MA, Pon CL
• Proteins from the prokaryotic nucleoid: primary and quaternary structure of the 15-kD Escherichia coli DNA binding protein H-NS.
• Mol Microbiol. 1988; 2: 323-9
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• The primary sequence of H-NS (136 amino acid residues, Mr = 15,402), an abundant Escherichia coli DNA-binding protein, has been elucidated and its quaternary structure has been investigated by protein-protein cross-linking reactions. It was found that H-NS exists predominantly as a dimer, even at very low concentrations, but may form tetramers at higher concentrations and that the protein-protein interaction responsible for the dimerization is chiefly hydrophobic.

• Kimura M, Kimura J, Davie P, Reinhardt R, Dijk J
• The amino acid sequence of a small DNA binding protein from the archaebacterium Sulfolobus solfataricus.
• FEBS Lett. 1984; 176: 176-8
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• The thermoacidophilic archaebacterium Sulfolobus solfataricus possesses several DNA binding proteins which may have a histone-like function. Two particularly dominant species have molecular masses of 7 and 10 kDa, respectively. We have purified one of the small proteins which occurs in relatively large amount and have determined its amino acid sequence. The protein is characterized by a high lysine content; in the N-terminal region the lysine residues occur in an alternating order: X-K-X-K-X-K-X-K. The amino acid sequence does not indicate any obvious homology to those DNA binding proteins whose sequences have been determined.

• Dijk J, White SW, Wilson KS, Appelt K
• On the DNA binding protein II from Bacillus stearothermophilus. I. Purification, studies in solution, and crystallization.
• J Biol Chem. 1983; 258: 4003-6
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• DNA binding protein II from Bacillus stearothermophilus has been purified as a single species from the nonribosomal cell fraction by a combination of gel filtration and ion exchange chromatography. The protein occurs in solution as a tetramer and is able to bind to 30 S, 50 S, and 70 S ribosomal particles. Circular dichroism studies show that the protein has approximately 45% alpha-helix. The secondary structure of the Bacillus protein is considerably more resistant to the effects of increasing temperature and urea concentration than the homologous protein (NS1 and NS2) from Escherichia coli. Proton magnetic resonance experiments show that the protein has a well folded, compact tertiary structure. The DNA binding protein has been crystallized from several precipitants as monoclinic needles and triclinic plates. The monoclinic form diffracts to at least 3.5 A and oscillation data from the native crystals have been collected. The protein is able to bind to both single- and double-stranded oligodeoxyribonucleotides. Upon binding, several changes occur in the protein NMR spectrum which may be used for further analysis of the mechanism of interaction.