An Amino Acid Region at the N-Terminus of Rat Hepatoma α1→2 Fucosyltransferase Modulates Enzyme Activity and Interaction with Lipids:  Strong Preference for Glycosphingolipids Containing Terminal Galβ1→3GalNAc- Structures†

Biochemistry ◽  
2001 ◽  
Vol 40 (19) ◽  
pp. 5708-5719 ◽  
Author(s):  
Anne L. Sherwood ◽  
Mark R. Stroud ◽  
Steven B. Levery ◽  
Eric H. Holmes
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Strong Preference ◽  
Amino Acid Region ◽  
N Terminus ◽  
Rat Hepatoma ◽  
Acid Region

scholarly journals Defining the Functionally Important Domain and Amino Acid Residues in Mycobacterium tuberculosis Integration Host Factor for Genome Stability, DNA Binding, and Integrative Recombination

2017 ◽  
Vol 199 (19) ◽  
Author(s):  
Narayanaswamy Sharadamma ◽  
Yadumurthy Harshavardhana ◽  
K. Muniyappa
Keyword(s):  
Dna Damage ◽  
Mycobacterium Tuberculosis ◽  
Amino Acid ◽  
Dna Binding ◽  
Integration Host Factor ◽  
Host Factor ◽  
Content Type ◽  
Amino Acid Region ◽  
N Terminus ◽  
Acid Region

ABSTRACT The integration host factor of Mycobacterium tuberculosis (mIHF) consists of a single polypeptide chain, the product of the ihf gene. We previously revealed that mIHF is a novel member of a new class of nucleoid-associated proteins that have important roles in DNA damage response, nucleoid compaction, and integrative recombination. The mIHF contains a region of 86 amino acids at its N terminus, absent from both α- and β-subunits of Escherichia coli IHF. However, the functional significance of an extra 86-amino-acid region in the full-length protein remains unknown. Here, we report the structure/function relationship of the DNA-binding and integrative recombination-stimulating activity of mIHF. Deletion mutagenesis showed that an extra 86-amino-acid region at the N terminus is dispensable; the C-terminal region possesses the sequences essential for its known biological functions, including the ability to suppress the sensitivity of E. coli ΔihfA and ΔihfB cells to DNA-damaging agents, DNA binding, DNA multimerization-circularization, and stimulation of phage L5 integrase-catalyzed integrative recombination. Single and double alanine substitutions at positions Arg170 and Arg171, located at the mIHF DNA-binding site, abrogated its capacity to suppress the sensitivity of E. coli ΔihfA and ΔihfB cells to DNA-damaging agents. The variants encoded by these mutant alleles failed to bind DNA and stimulate integrative recombination. Interestingly, the DNA-binding activity of the mIHF-R173A variant remained largely unaffected; however, it was unable to stimulate integrative recombination, thus revealing a separation-of-function allele of mIHF. The functional and structural characterization of this separation-of-function allele of mIHF could reveal previously unknown functions of IHF. IMPORTANCE The integration host factor of Mycobacterium tuberculosis is a novel nucleoid-associated protein. mIHF plays a vital role in DNA damage response, nucleoid compaction, and integrative recombination. Intriguingly, mIHF contains an extra 86-amino-acid region at its N terminus, absent from both α- and β-subunits of Escherichia coli IHF, whose functional significance is unknown. Furthermore, a triad of arginine residues located at the mIHF-DNA interface have been implicated in a range of its functions. Here, we reveal the roles of N- and C-terminal regions of mIHF and the individual residues in the Arg triad for their ability to provide protection in vivo against DNA damage, bind DNA, and stimulate integrase-catalyzed site-specific recombination.

scholarly journals A Carboxy-Terminal 16-Amino-Acid Region of ς38 ofEscherichia coli Is Important for Transcription under High-Salt Conditions and Sigma Activities In Vivo

2000 ◽  
Vol 182 (16) ◽  
pp. 4628-4631 ◽  
Author(s):  
Mio Ohnuma ◽  
Nobuyuki Fujita ◽  
Akira Ishihama ◽  
Kan Tanaka ◽  
Hideo Takahashi
Keyword(s):  
Amino Acid ◽  
Bacterial Species ◽  
High Potassium ◽  
Transcription Analysis ◽  
Amino Acid Region ◽  
Sigma Subunit ◽  
Carboxy Terminal ◽  
Acid Region

ABSTRACT ς38 (or ςS, the rpoS gene product) is a sigma subunit of RNA polymerase in Escherichia coli and directs transcription from a number of stationary-phase promoters as well as osmotically inducible promoters. In this study, we analyzed the function of the carboxy-terminal 16-amino-acid region of ς38 (residues 315 to 330), which is well conserved among the rpoS gene products of enteric bacterial species. Truncation of this region was shown to result in the loss of sigma activity in vivo using promoter-lacZ fusion constructs, but the mutant ς38 retained the binding activity in vivo to the core enzyme. The in vitro transcription analysis revealed that the transcription activity of ς38 holoenzyme under high potassium glutamate concentrations was significantly decreased by the truncation of the carboxy-terminal tail element.

scholarly journals The DNA-binding specificity of the hepatocyte nuclear factor 3/forkhead domain is influenced by amino-acid residues adjacent to the recognition helix.

1994 ◽  
Vol 14 (4) ◽  
pp. 2755-2766 ◽  
Author(s):  
D G Overdier ◽  
A Porcella ◽  
R H Costa
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Hepatocyte Nuclear Factor ◽  
Binding Properties ◽  
Winged Helix ◽  
Amino Acid Region ◽  
Dna Binding Specificity ◽  
Recognition Helix ◽  
Dna Binding Properties ◽  
Acid Region

Three distinct hepatocyte nuclear factor 3 (HNF-3) proteins (HNF-3 alpha, -3 beta, and -3 gamma) are known to regulate the transcription of liver-specific genes. The HNF-3 proteins bind to DNA as a monomer through a modified helix-turn-helix, known as the winged helix motif, which is also utilized by a number of developmental regulators, including the Drosophila homeotic forkhead (fkh) protein. We have previously described the isolation, from rodent tissue, of an extensive family of tissue-specific HNF-3/fkh homolog (HFH) genes sharing homology in their winged helix motifs. In this report, we have determined the preferred DNA-binding consensus sequence for the HNF-3 beta protein as well as for two divergent family members, HFH-1 and HFH-2. We show that these HNF-3/fkh proteins bind to distinct DNA sites and that the specificity of protein recognition is dependent on subtle nucleotide alterations in the site. The HNF-3, HFH-1, and HFH-2 consensus binding sequences were also used to search DNA regulatory regions to identify potential target genes. Furthermore, an analysis of the DNA-binding properties of a series of HFH-1/HNF-3 beta protein chimeras has allowed us to identify a 20-amino-acid region, located adjacent to the DNA recognition helix, which contributes to DNA-binding specificity. These sequences are not involved in base-specific contacts and include residues which diverge within the HNF-3/fkh family. Replacement of this 20-amino-acid region in HNF-3 beta with corresponding residues from HFH-1 enabled the HNF-3 beta recognition helix to bind only HFH-1-specific DNA-binding sites. We propose a model in which this 20-amino-acid flanking region influences the DNA-binding properties of the recognition helix.

scholarly journals Rho1p-Bni1p-Spa2p Interactions: Implication in Localization of Bni1p at the Bud Site and Regulation of the Actin Cytoskeleton inSaccharomyces cerevisiae

1998 ◽  
Vol 9 (5) ◽  
pp. 1221-1233 ◽  
Author(s):  
Takeshi Fujiwara ◽  
Kazuma Tanaka ◽  
Akihisa Mino ◽  
Mitsuhiro Kikyo ◽  
Kazuo Takahashi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Actin Cytoskeleton ◽  
Binding Protein ◽  
Microscopic Analysis ◽  
Mitogen Activated Protein Kinase ◽  
Gene Encoding ◽  
Amino Acid Region ◽  
Genes Encoding ◽  
Mitogen Activated Protein ◽  
Acid Region

Rho1p is a yeast homolog of mammalian RhoA small GTP-binding protein. Rho1p is localized at the growth sites and required for bud formation. We have recently shown that Bni1p is a potential target of Rho1p and that Bni1p regulates reorganization of the actin cytoskeleton through interactions with profilin, an actin monomer-binding protein. Using the yeast two-hybrid screening system, we cloned a gene encoding a protein that interacted with Bni1p. This protein, Spa2p, was known to be localized at the bud tip and to be implicated in the establishment of cell polarity. The C-terminal 254 amino acid region of Spa2p, Spa2p(1213–1466), directly bound to a 162-amino acid region of Bni1p, Bni1p(826–987). Genetic analyses revealed that both thebni1 and spa2 mutations showed synthetic lethal interactions with mutations in the genes encoding components of the Pkc1p-mitogen-activated protein kinase pathway, in which Pkc1p is another target of Rho1p. Immunofluorescence microscopic analysis showed that Bni1p was localized at the bud tip in wild-type cells. However, in the spa2 mutant, Bni1p was not localized at the bud tip and instead localized diffusely in the cytoplasm. A mutant Bni1p, which lacked the Rho1p-binding region, also failed to be localized at the bud tip. These results indicate that both Rho1p and Spa2p are involved in the localization of Bni1p at the growth sites where Rho1p regulates reorganization of the actin cytoskeleton through Bni1p.

The F12-Vif derivative Chim3 inhibits HIV-1 replication in CD4+ T lymphocytes and CD34+-derived macrophages by blocking HIV-1 DNA integration

Blood ◽  
2009 ◽  
Vol 113 (15) ◽  
pp. 3443-3452 ◽  
Author(s):  
Simona Porcellini ◽  
Luca Alberici ◽  
Francesco Gubinelli ◽  
Rossella Lupo ◽  
Clelia Olgiati ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Amino Acid ◽  
T Lymphocytes ◽  
Genetic Manipulation ◽  
Natural Resistance ◽  
Dna Integration ◽  
Amino Acid Region ◽  
Antiviral Function ◽  
Hiv 1 ◽  
Acid Region

Abstract The viral infectivity factor (Vif) is essential for HIV-1 infectivity and hence is an ideal target for promising anti–HIV-1/AIDS gene therapy. We previously demonstrated that F12-Vif mutant inhibits HIV-1 replication in CD4+ T lymphocytes. Despite macrophage relevance to HIV-1 pathogenesis, most gene therapy studies do not investigate macrophages because of their natural resistance to genetic manipulation. Here, we confirm the F12-Vif antiviral activity also in macrophages differentiated in vitro from transduced CD34+ human stem cells (HSCs). Moreover, we identified the 126- to 170-amino-acid region in the C-terminal half of F12-Vif as responsible for its antiviral function. Indeed, Chim3 protein, containing this 45-amino-acid region embedded in a WT-Vif backbone, is as lethal as F12-Vif against HIV-1. Of major relevance, we demonstrated a dual mechanism of action for Chim3. First, Chim3 functions as a transdominant factor that preserves the antiviral function of the natural restriction factor APOBEC3G (hA3G). Second, Chim3 blocks the early HIV-1 retrotranscript accumulation and thereby HIV-1 DNA integration regardless of the presence of WT-Vif and hA3G. In conclusion, by impairing the early steps of HIV-1 life cycle, Chim3 conceivably endows engineered cells with survival advantage, which is required for the efficient immune reconstitution of patients living with HIV/AIDS.

Identification of an amino acid region supporting specific methionyl-tRNA synthetase: tRNA recognition

1989 ◽  
Vol 208 (3) ◽  
pp. 429-443 ◽  
Author(s):  
Patrice Mellot ◽  
Yves Mechulam ◽  
Daniel Le Corre ◽  
Sylvain Blanquet ◽  
Guy Fayat
Keyword(s):  
Amino Acid ◽  
Trna Synthetase ◽  
Trna Recognition ◽  
Amino Acid Region ◽  
Methionyl Trna Synthetase ◽  
Acid Region

scholarly journals Contribution of Amino Acid Region 334−335 from Factor Va Heavy Chain to the Catalytic Efficiency of Prothrombinase†

Biochemistry ◽  
2008 ◽  
Vol 47 (26) ◽  
pp. 6840-6850 ◽  
Author(s):  
Melissa A. Barhoover ◽  
Tivadar Orban ◽  
Daniel O. Beck ◽  
Michael A. Bukys ◽  
Michael Kalafatis
Keyword(s):  
Amino Acid ◽  
Heavy Chain ◽  
Catalytic Efficiency ◽  
Amino Acid Region ◽  
Factor Va ◽  
Acid Region

scholarly journals A divergent CheW confers plasticity to nucleoid-associated chemosensory arrays

2017 ◽  
Author(s):  
Annick Guiseppi ◽  
Juan Jesus Vicente ◽  
Julien Herrou ◽  
Deborah Byrne ◽  
Aurelie Barneoud ◽  
...  
Keyword(s):  
Amino Acid ◽  
Histidine Kinase ◽  
Environmental Changes ◽  
Myxococcus Xanthus ◽  
Accessory Protein ◽  
Cellular Functions ◽  
The Core ◽  
Amino Acid Region ◽  
Chemosensory Systems ◽  
Acid Region

ABSTRACTChemosensory systems are highly organized signaling pathways that allow bacteria to adapt to environmental changes. The Frz chemosensory system from M. xanthus possesses two CheW-like proteins, FrzA (the core CheW) and FrzB. We found that FrzB does not interact with FrzE (the cognate CheA) as it lacks the amino acid region responsible for this interaction. FrzB, instead, acts upstream of FrzCD in the regulation of M. xanthus chemotaxis behaviors and activates the Frz pathway by allowing the formation and distribution of multiple chemosensory clusters on the nucleoid. These results, together, show that the lack of the CheA-interacting region in FrzB confers new functions to this small protein.AUTHOR SUMMARYChemosensory systems are signaling complexes that are widespread in bacteria and allow the modulation of different cellular functions, such as taxis and development, in response to the environment. We show that the Myxococcus xanthus FrzB is a divergent CheW lacking the region involved in the interaction with the histidine kinase FrzE. Instead, it acts upstream of FrzCD to allow the formation of multiple distributed Frz chemosensory arrays at the nucleoid. The loss of the CheA-interacting region in FrzB might have been selected to confer plasticity to nucleoid-associated chemosensory systems. By unraveling a new accessory protein and its function, this work opens new insights into the knowledge of the regulatory potentials of bacterial chemosensory systems.

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