Phage integrases: biology and applications.

https://www.cell.com/cell/pdf/S0092-8674(00)80787-4.pdf

Polypeptide flux through bacterial Hsp70: DnaK cooperates with trigger factor in chaperoning nascent chains.

Publisher Summary This chapter deals with the structure of the molten globules of various globular proteins revealed by the recent experimental studies. Recent advances in experimental techniques, including hydrogen-exchange NMR, solution X-ray scattering, and protein engineering, have provided detailed pictures of the molten globules for these proteins. The molten globule state has heterogeneous structures, in which one portion of a molecule is more organized and native-like with the other portions being less organized, although the overall structure satisfies the criteria of the molten globule state (compactness, the presence of secondary structure, and the lack of rigid tertiary structure). The chapter describes how the molten globule state has been identified as the intermediate of kinetic refolding and discusses the kinetic roles of the molten globule state in protein folding. The chapter also discusses thermodynamic stability and cooperativity of the molten globule state from the viewpoint of the hierarchy of protein folding, in which the molten globule state plays a role as a junction of two levels of the hierarchy.

Role of the molten globule state in protein folding.

Mycobacterium tuberculosis Invasion of Macrophages: Linking Bacterial Gene Expression to Environmental Cues

http://archive.gersteinlab.org/papers/e-print/ribotunnel/preprint.pdf

The geometry of the ribosomal polypeptide exit tunnel.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/j.febslet.2006.09.004

Transcriptional autoregulation by Mycobacterium tuberculosis PhoP involves recognition of novel direct repeat sequences in the regulatory region of the promoter

The integrase protein (Int) from bacteriophage λ catalyzes the insertion and excision of the viral genome into and out of Escherichia coli. It is a member of the λ-Int family of site-specific recombinases that catalyze a diverse array of DNA rearrangements in archaebacteria, eubacteria, and yeast and belongs to the subset of this family that possesses two autonomous DNA-binding domains. The heterobivalent properties of Int can be decomposed into a carboxyl-terminal domain that executes the DNA cleavage and ligation reactions and a smaller amino-terminal domain that binds to an array of conserved DNA sites within the phage arms, thereby arranging Int protomers within the higher-order recombinogenic complex. We have determined that residues Met-1 to Leu-64 of Int constitute the minimal arm-type DNA-binding domain (INT-DBD1–64) and solved the solution structure by using NMR. We show that the INT-DBD1–64 is a novel member of the growing family of three-stranded β-sheet DNA-binding proteins, because it supplements this motif with a disordered amino-terminal basic tail that is important for arm-site binding. A model of the arm-DNA-binding domain recognizing its cognate DNA site is proposed on the basis of similarities with the analogous domain of Tn916 Int and is discussed in relation to other features of the protein.

Arm-site binding by λ-integrase: Solution structure and functional characterization of its amino-terminal domain

λ Integrase (Int) has the distinctive ability to bridge two different and well separated DNA sequences. This heterobivalent DNA binding is facilitated by accessory DNA bending proteins that bring flanking Int sites into proximity. The regulation of λ recombination has long been perceived as a structural phenomenon based upon the accessory protein-dependent Int bridges between high-affinity arm-type (bound by the small N-terminal domain) and low-affinity core-type DNA sites (bound by the large C-terminal domain). We show here that the N-terminal domain is not merely a guide for the proper positioning of Int protomers, but is also a context-sensitive modulator of recombinase functions. In full-length Int, it inhibits C-terminal domain binding and cleavage at the core sites. Surprisingly, its presence as a separate molecule stimulates the C-terminal domain functions. The inhibition in full-length Int is reversed or overcome in the presence of arm-type oligonucleotides, which form specific complexes with Int and core-type DNA. We consider how these results might influence models and experiments pertaining to the large family of heterobivalent recombinases.

https://www.embopress.org/doi/pdf/10.1093/emboj/20.5.1203

The small DNA binding domain of λ integrase is a context‐sensitive modulator of recombinase functions

The ability to sense acid stress and mount an appropriate adaptive response by Mycobacterium tuberculosis, which adapts a long-term residence in the macrophage phagosome, remains one of the critical features that defines mycobacterial physiology and its intracellular location. To understand the mechanistic basis of adaptation of the intracellular pathogen, we studied global regulation of M. tuberculosis gene expression in response to acid stress. Although recent studies indicate a role for the virulence-associated phoP locus in pH-driven adaptation, in this study, we discovered a strikingly novel regulatory mechanism, which controls acid-stress homeostasis. Using mycobacterial protein fragment complementation and in vitro interaction analyses, we demonstrate that PhoP interacts with acid-inducible extracytoplasmic SigE (one of the 13 M. tuberculosis sigma factors) to regulate a complex transcriptional program. Based on these results, we propose a model to suggest that PhoP-SigE interaction represents a major requirement for the global acid stress response, absence of which leads to strongly reduced survival of the bacilli under acidic pH conditions. These results account for the significant growth attenuation of the phoP mutant in both cellular and animal models, and unravel the underlying global mechanism of how PhoP induces an adaptive program in response to acid stress.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/mmi.13635

Mycobacterium tuberculosis virulence‐regulator PhoP interacts with alternative sigma factor SigE during acid‐stress response

Mycobacterium tuberculosis PhoP regulates the expression of unknown virulence determinants and the biosynthesis of complex lipids. PhoP, like other members of the OmpR family, comprises a phosphorylation domain at the amino-terminal half and a DNA-binding domain at the carboxy-terminal half of the protein. To explore structural effect of protein phosphorylation and to examine effect of phosphorylation on DNA binding, purified PhoP was phosphorylated by acetyl phosphate in a reaction that was dependent on Mg2+ and Asp-71. Protein phosphorylation was not required for DNA binding; however, phosphorylation enhanced in vitro DNA binding through protein-protein interaction(s). Evidence is presented here that the protein-protein interface is different in the unphosphorylated and phosphorylated forms of PhoP and that specific DNA binding plays a critical role in changing the nature of the protein-protein interface. We show that phosphorylation switches the transactivation domain to a different conformation, which specifies additional protein-protein contacts between PhoP protomers bound to adjacent cognate sites. Together, our observations raise the possibility that PhoP, in the unphosphorylated and phosphorylated forms, may be capable of adopting different orientations as it binds to a vast array of genes to activate or repress transcription.

Dibyendu Sarkar

Papers

Transcriptional autoregulation by Mycobacterium tuberculosis PhoP involves recognition of novel direct repeat sequences in the regulatory region of the promoter

Arm-site binding by λ-integrase: Solution structure and functional characterization of its amino-terminal domain

The small DNA binding domain of λ integrase is a context‐sensitive modulator of recombinase functions

Mycobacterium tuberculosis virulence‐regulator PhoP interacts with alternative sigma factor SigE during acid‐stress response

PhoP-PhoP Interaction at Adjacent PhoP Binding Sites Is Influenced by Protein Phosphorylation