Solution Structure of the B-Myb DNA-Binding Domain:  A Possible Role for Conformational Instability of the Protein in DNA Binding and Control of Gene Expression<sup>,</sup>

McIntosh, Pauline B.; Frenkiel, Thomas A.; Wollborn, Ute; McCormick, John E.; Klempnauer, K H; Feeney, J.; Carr, Mark D.

doi:10.1021/bi972861z

Cited by 24 publications

(22 citation statements)

References 35 publications

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“…At the end of the procedure (cycle 7) unique assignments were obtained for 94.3% (1498/ 1589) of the NOE peaks picked in the 15 N/ 1 H NOESY-HSQC spectrum and 96.3% (4030/4184) of those identified in the 13 C/ 1 H HMQC-NOESY spectrum. This level of success compares very favorably with our experience of manual iterative assignment of NOE peaks in protein spectra (42,43,47), and the automatic approach took only a few days as compared with several months for manual assignment. The uniquely assigned NOE peaks produced 2845 non-redundant 1 H to 1 H upper distance limits, which were used as the principle constraints in the final rounds of structural calculations.…”

Section: Resultssupporting

confidence: 64%

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

Carr

Bloemink

Dentten

et al. 2003

Journal of Biological Chemistry

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The closely related mycobacteria responsible for tuberculosis produce an unusually high number of secreted proteins, many of which are clearly implicated in pathogenesis and protective immunity. Falling within this category are the closely related proteins MPB70 and MPB83. The structure of MPB70 reveals a complex and novel bacterial fold, which has clear structural homology to the two C-terminal FAS1 domains of the cell adhesion protein fasciclin I, whose structures were reported very recently. Assessment of the surface features of MPB70, the sequence divergence between MPB70 and MPB83, the conservation of residues across a group of FAS1 domains, and the locations of disease-inducing mutations in ␤ig-h3 strongly suggests that MPB70 and MPB83 contain two functional surfaces on opposite faces, which are probably involved in binding to host cell proteins. This analysis also suggests that these functional surfaces are retained in the FAS1 proteins associated with mediating interactions between cells and the extracellular matrix (fasciclin I, periostin, and ␤ig-h3) and furthermore that some of the human corneal disease-inducing substitutions identified in ␤ig-h3 will perturb interactions at these sites.Tuberculosis remains one of the most significant infectious diseases of humans, with about one-third of the world's population currently infected resulting in about 3 million deaths annually (1, 2). The bacteria responsible for tuberculosis belong to the Mycobacterium tuberculosis complex, which is a group of highly related mycobacteria. The complex includes M. tuberculosis, which causes the majority of human tuberculosis, and Mycobacterium bovis, which leads to tuberculosis in a range of domesticated and wild animals. Analysis of the complete M. tuberculosis H37Rv genome has identified the genes for 4006 proteins (3, 4), however, it has so far proved possible to assign specific functions to only about half of the predicted proteins. In addition, we still have relatively little information about which M. tuberculosis complex proteins are essential for pathogenesis, or associated with the stimulation of protective immunity, and even less knowledge of their structures, functions, and mechanisms of action. The finding that only live mycobacterial vaccines provide significant protection against tuberculosis infection (5) clearly indicates a key role for secreted M. tuberculosis complex proteins in stimulating protective immunity and highlights the importance of investigating major secreted antigenic proteins such as MPB70, MPB83, ESAT-6, and CFP-10 (6 -10).The mature M. bovis protein MPB70 and its identical M. tuberculosis homologue MPT70 (Rv2875) are stable, 163-residue polypeptides, that are efficiently secreted from mycobacterial cells following cleavage of a 30-residue signal peptide (3, 6). The proteins contain a single disulfide bond linking Cys-8 and Cys-142 but show no other form of post-translational modification. MPB70 is a major serodominant antigen of M. bovis, which also stimulates cellular immune responses du...

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Section: Resultssupporting

confidence: 64%

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

Carr

Bloemink

Dentten

et al. 2003

Journal of Biological Chemistry

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“…This property has been reported for a number of protein and nucleic acid binding sites on proteins (31,32) and may be an important factor controlling the specificity of the binding interaction. It has been shown by Kay and co-workers (33,34) that the flexible region of the peptide binding site in the phospholipase C-␥1 SH2 domain has a much wider ligand binding specificity than the more rigid phosphotyrosine binding site.…”

Section: Backbone Mobility Of the Binding Site For N-mmp-3-thementioning

confidence: 68%

“…1C 32 -Asn 33 in the AB ␤-hairpin were also marginally above the threshold (mean plus 1 standard deviation) considered to be significant (Fig. 1C).…”

Section: Resultsmentioning

confidence: 84%

The Effect of Matrix Metalloproteinase Complex Formation on the Conformational Mobility of Tissue Inhibitor of Metalloproteinases-2 (TIMP-2)

Williamson

Muskett

Howard

et al. 1999

Journal of Biological Chemistry

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The backbone mobility of the N-terminal domain of tissue inhibitor of metalloproteinases-2 (N-TIMP-2) was determined both for the free protein and when bound to the catalytic domain of matrix metalloproteinase-3 (N-MMP-3). Regions of the protein with internal motion were identified by comparison of the T 1 and T 2 relaxation times and 1 H-15 N nuclear Overhauser effect values for the backbone amide 15 N signals for each residue in the sequence. This analysis revealed rapid internal motion on the picosecond to nanosecond time scale for several regions of free N-TIMP-2, including the extended ␤-hairpin between ␤-strands A and B, which forms part of the MMP binding site. Evidence of relatively slow motion indicative of exchange between two or more local conformations on a microsecond to millisecond time scale was also found in the free protein, including two other regions of the MMP binding site (the CD and EF loops). On formation of a tight N-TIMP-2⅐N-MMP-3 complex, the rapid internal motion of the AB ␤-hairpin was largely abolished, a change consistent with tight binding of this region to the MMP-3 catalytic domain. The extended AB ␤-hairpin is not a feature of all members of the TIMP family; therefore, the binding of this highly mobile region to a site distant from the catalytic cleft of the MMPs suggests a key role in TIMP-2 binding specificity.Breakdown of the extracellular matrix is an important event in many normal and pathological processes, such as growth, wound repair, tumor metastasis, and arthritis (1-3). A large family of zinc-dependent proteinases, the matrix metalloproteinases (MMPs), 1 are thought to be primarily responsible for this matrix catabolism. The activities of the MMP family in the extracellular matrix are highly regulated by transcriptional control, zymogen activation, and inhibition by a family of specific protein inhibitors, the tissue inhibitors of metalloproteinases or TIMPs (4). The TIMPs bind tightly to the active proteinases to form an inactive TIMP⅐MMP complex (5). Four mammalian TIMP proteins have now been identified (TIMP-1 to -4) (6 -9), and their high degree of sequence similarity and conservation of 12 Cys residues suggests that each consists of the same basic fold but with some variations in loop structures and glycosylation. The location of the MMP inhibitory site on the TIMP molecule has been shown to reside predominantly in the N-terminal two-thirds of the protein, defined by the first three disulfide bonds. This domain (N-TIMP) can be expressed independently to generate a fully folded, stable, and active inhibitor (10, 11).High resolution three-dimensional structures are now available for both the active N-terminal domain of TIMP-2 (12) and for the full-length inhibitor (13). Crystal structures have also been published for full-length TIMP-1 and TIMP-2 in complexes with the catalytic domains of MMP-3 (N-MMP-3) and MT1-MMP, respectively (14, 15). The structures of the TIMP⅐MMP complexes, together with NMR data on chemical shift perturbation seen for N-TIMP-2 on complex fo...

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“…Interestingly, in vitro translated full-length B-Myb did not bind to Myb binding site in the SV40 promoter, but the C-terminal truncation mutants of BMyb or B-Myb DNA binding domain fused to c-Myb were able to bind the same sequences suggesting that C-terminus of B-Myb exerts additional limitation to DNA binding . Third, the NMR structure for the DNA binding domain of B-Myb revealed that despite extensive sequence similarity in the R2 and R3 regions, the C-terminal region of BMyb R2 shows a poorly de®ned structure, re¯ecting the existence of multiple conformations in slow to intermediate exchange (McIntosh et al, 1998). This contrasts with the tertiary structure reported for c-Myb R2R3, in which both R2 and R3 have the same folding and the C-terminal region of R2 forms a stable, wellde®ned helix (Ogata et al, 1994).…”

Section: Sequence Recognition By Myb Dna Binding Motifmentioning

confidence: 99%

The myb gene family in cell growth, differentiation and apoptosis

1999

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The myb gene family consists of three members, named A, B and c-myb which encode nuclear proteins that function as transcriptional transactivators. Proteins encoded by these three genes exhibit a tripartate structure with an N-terminal DNA-binding domain, a central transactivation domain and a C-terminal regulatory domain. These proteins exhibit highest homology in their DNA binding domains and appear to bind DNA with overlapping sequence speci®cities. Transactivation by myb gene family varies considerably depending on cell type and promoter context suggesting a dependence on interaction with other cell type speci®c co-factors. While the C-terminal domains of A-Myb and c-Myb proteins exert a negative regulatory eect on their transcriptional transactivation function, the C-terminal domain of BMyb appears to function as a positive regulator of this activity. One or more of these proteins interact with other transcription factors such as Ets-2, CEBP and NF-M. In addition, expression of these genes is cell cycleregulated and inhibition of their expression with antisense oligonucleotides has been found to aect cell cycleprogression, cell division and/or dierentiation. Members of the myb gene family exhibit dierent temporal and spatial expression patterns suggesting a distinctive function for each of these genes. Gene knockout experiments show that these genes play an essential role in development. Loss of c-myb function results in embryonic lethality due to failure of fetal hepatic hematopoiesis. A-myb null mutant mice, on the other hand are viable but exhibit growth abnormalities, and defects in spermatogenesis and female breast development. While the role of c-myb in oncogenesis is well established, future experiments are likely to provide further clues regarding the role of A-myb and B-myb in tumorigenesis.

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Solution Structure of the B-Myb DNA-Binding Domain: A Possible Role for Conformational Instability of the Protein in DNA Binding and Control of Gene Expression^,

Cited by 24 publications

References 35 publications

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

The Effect of Matrix Metalloproteinase Complex Formation on the Conformational Mobility of Tissue Inhibitor of Metalloproteinases-2 (TIMP-2)

The myb gene family in cell growth, differentiation and apoptosis

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Solution Structure of the B-Myb DNA-Binding Domain: A Possible Role for Conformational Instability of the Protein in DNA Binding and Control of Gene Expression,

Cited by 24 publications

References 35 publications

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

Solution Structure of the Mycobacterium tuberculosis Complex Protein MPB70

The Effect of Matrix Metalloproteinase Complex Formation on the Conformational Mobility of Tissue Inhibitor of Metalloproteinases-2 (TIMP-2)

The myb gene family in cell growth, differentiation and apoptosis

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Product

Resources

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Solution Structure of the B-Myb DNA-Binding Domain: A Possible Role for Conformational Instability of the Protein in DNA Binding and Control of Gene Expression^,