scholarly journals The BBA01 Protein, a Member of Paralog Family 48 from Borrelia burgdorferi, Is Potentially Interchangeable with the Channel-Forming Protein P13

2006 ◽  
Vol 188 (12) ◽  
pp. 4207-4217 ◽  
Author(s):  
Marija Pinne ◽  
Katrin Denker ◽  
Elin Nilsson ◽  
Roland Benz ◽  
Sven Bergström
Keyword(s):  
Borrelia Burgdorferi ◽  
Single Channel ◽  
Sequence Similarity ◽  
Shuttle Vector ◽  
Protein A ◽  
Gene Families ◽  
Wild Type ◽  
C Terminus ◽  
High Degree ◽  
Paralog Family

ABSTRACT The Borrelia burgdorferi genome exhibits redundancy, with many plasmid-carried genes belonging to paralogous gene families. It has been suggested that certain paralogs may be necessary in various environments and that they are differentially expressed in response to different conditions. The chromosomally located p13 gene which codes for a channel-forming protein belongs to paralog family 48, which consists of eight additional genes. Of the paralogous genes from family 48, the BBA01 gene has the highest homology to p13. Herein, we have inactivated the BBA01 gene in B. burgdorferi strain B31-A. This mutant shows no apparent phenotypic difference compared to the wild type. However, analysis of BBA01 in a C-terminal protease A (CtpA)-deficient background revealed that like P13, BBA01 is posttranslationally processed at its C terminus. Elevated BBA01 expression was obtained in strains with the BBA01 gene introduced on the shuttle vector compared to the wild-type strain. We could further demonstrate that BBA01 is a channel-forming protein with properties surprisingly similar to those of P13. The single-channel conductance, of about 3.5 nS, formed by BBA01 is comparable to that of P13, which together with the high degree of sequence similarity suggests that the two proteins may have similar and interchangeable functions. This is further strengthened by the up-regulation of the BBA01 protein and its possible localization in the outer membrane in a p13 knockout strain, thus suggesting that P13 can be replaced by BBA01.

scholarly journals The TFIID Components Human TAFII140 andDrosophila BIP2 (TAFII155) Are Novel Metazoan Homologues of Yeast TAFII47 Containing a Histone Fold and a PHD Finger

2001 ◽  
Vol 21 (15) ◽  
pp. 5109-5121 ◽  
Author(s):  
Yann-Gaël Gangloff ◽  
Jean-Christophe Pointud ◽  
Sylvie Thuault ◽  
Lucie Carré ◽  
Christophe Romier ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Sequence Similarity ◽  
Protein A ◽  
Amino Acid Sequences ◽  
Molecular Organization ◽  
Phd Finger ◽  
Histone Fold ◽  
Histone Fold Domain ◽  
High Degree

ABSTRACT The RNA polymerase II transcription factor TFIID comprises the TATA binding protein (TBP) and a set of TBP-associated factors (TAFIIs). TFIID has been extensively characterized for yeast, Drosophila, and humans, demonstrating a high degree of conservation of both the amino acid sequences of the constituent TAFIIs and overall molecular organization. In recent years, it has been assumed that all the metazoan TAFIIs have been identified, yet no metazoan homologues of yeast TAFII47 (yTAFII47) and yTAFII65 are known. Both of these yTAFIIs contain a histone fold domain (HFD) which selectively heterodimerizes with that of yTAFII25. We have cloned a novel mouse protein, TAFII140, containing an HFD and a plant homeodomain (PHD) finger, which we demonstrated by immunoprecipitation to be a mammalian TFIID component. TAFII140 shows extensive sequence similarity toDrosophila BIP2 (dBIP2) (dTAFII155), which we also show to be a component of DrosophilaTFIID. These proteins are metazoan homologues of yTAFII47 as their HFDs selectively heterodimerize with dTAFII24 and human TAFII30, metazoan homologues of yTAFII25. We further show that yTAFII65 shares two domains with theDrosophila Prodos protein, a recently described potential dTAFII. These conserved domains are critical for yTAFII65 function in vivo. Our results therefore identify metazoan homologues of yTAFII47 and yTAFII65.

scholarly journals Oligopeptide Permease A5 Modulates Vertebrate Host-Specific Adaptation of Borrelia burgdorferi

2011 ◽  
Vol 79 (8) ◽  
pp. 3407-3420 ◽  
Author(s):  
B. V. Subba Raju ◽  
Maria D. Esteve-Gassent ◽  
S. L. Rajasekhar Karna ◽  
Christine L. Miller ◽  
Tricia A. Van Laar ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Shuttle Vector ◽  
Immunoblot Analysis ◽  
Vertebrate Host ◽  
Transcriptional Analysis ◽  
Wild Type ◽  
Content Type ◽  
Specific Adaptation ◽  
Protein Levels ◽  
Oligopeptide Permease

ABSTRACTBorrelia burgdorferi, the agent of Lyme disease, undergoes rapid adaptive gene expression in response to signals unique to its arthropod vector or vertebrate hosts. Among the upregulated genes under vertebrate host conditions is one of the five annotated homologs of oligopeptide permease A (OppA5, BBA34). A mutant lackingoppA5was constructed in an lp25-deficient isolate ofB. burgdorferistrain B31, and the minimal regions of infectivity were restored via a shuttle vector pBBE22 with or without an intact copy ofbba34. Immunoblot analysis of thebba34mutant revealed a reduction in the levels of RpoS, BosR, and CsrABbwith a concomitant reduction in the levels of OspC, DbpA, BBK32, and BBA64. There were no changes in the levels of OspA, NapA, P66, and three other OppA orthologs. Quantitative transcriptional analysis correlated with the changes in the protein levels. However, thebba34mutant displayed comparable infectivities in the C3H/HeN mice and the wild-type strain, despite the reduction in several pathogenesis-related proteins. Supplementation of the growth medium with increased levels of select components, notably sodium acetate and sodium bicarbonate, restored the levels of several proteins in thebba34mutant to wild-type levels. We speculate that the transport of acetate appears to contribute to the accumulation of key metabolites, like acetyl phosphate, that facilitate the adaptation ofB. burgdorferito the vertebrate host by the activation of the Rrp2-RpoN-RpoS pathway. These studies underscore the importance of solute transport to host-specific adaptation ofB. burgdorferi.

scholarly journals pbpB, a Gene Coding for a Putative Penicillin-Binding Protein, Is Required for Aerobic Nitrogen Fixation in the Cyanobacterium Anabaena sp. Strain PCC7120

2001 ◽  
Vol 183 (2) ◽  
pp. 628-636 ◽  
Author(s):  
Sara Lázaro ◽  
Francisca Fernández-Piñas ◽  
Eduardo Fernández-Valiente ◽  
Amaya Blanco-Rivero ◽  
Francisco Leganés
Keyword(s):  
Sequence Similarity ◽  
Shuttle Vector ◽  
Molecular Size ◽  
Nitrogen Deprivation ◽  
Wild Type ◽  
Reading Frame ◽  
Cyanobacterium Anabaena ◽  
In The Wild ◽  
Peptidoglycan Layer

ABSTRACT Transposon mutagenesis of Anabaena sp. strain PCC7120 led to the isolation of a mutant strain, SNa1, which is unable to fix nitrogen aerobically but is perfectly able to grow with combined nitrogen (i.e., nitrate). Reconstruction of the transposon mutation of SNa1 in the wild-type strain reproduced the phenotype of the original mutant. The transposon had inserted within an open reading frame whose translation product shows significant homology with a family of proteins known as high-molecular-weight penicillin-binding proteins (PBPs), which are involved in the synthesis of the peptidoglycan layer of the cell wall. A sequence similarity search allowed us to identify at least 12 putative PBPs in the recently sequencedAnabaena sp. strain PCC7120 genome, which we have named and organized according to predicted molecular size and theEscherichia coli nomenclature for PBPs; based on this nomenclature, we have denoted the gene interrupted in SNal aspbpB and its product as PBP2. The wild-type form ofpbpB on a shuttle vector successfully complemented the mutation in SNa1. In vivo expression studies indicated that PBP2 is probably present when both sources of nitrogen, nitrate and N2, are used. When nitrate is used, the function of PBP2 either is dispensable or may be substituted by other PBPs; however, under nitrogen deprivation, where the differentiation of the heterocyst takes place, the role of PBP2 in the formation and/or maintenance of the peptidoglycan layer is essential.

scholarly journals The ς70 Transcription Factor TyrR Has Zinc-Stimulated Phosphatase Activity That Is Inhibited by ATP and Tyrosine

2000 ◽  
Vol 182 (4) ◽  
pp. 1053-1061 ◽  
Author(s):  
Shimin Zhao ◽  
Qin Zhu ◽  
Ronald L. Somerville
Keyword(s):  
Amino Acid ◽  
Phosphatase Activity ◽  
Sequence Similarity ◽  
Zinc Ion ◽  
Amino Acid Residues ◽  
Protein Residues ◽  
E Coli ◽  
C Terminus ◽  
Atp Analogues ◽  
High Degree

ABSTRACT The TyrR protein of Escherichia coli (513 amino acid residues) is the chief transcriptional regulator of a group of genes that are essential for aromatic amino acid biosynthesis and transport. The TyrR protein can function either as a repressor or as an activator. The central region of the TyrR protein (residues 207 to 425) is similar to corresponding polypeptide segments of the NtrC protein superfamily. Like the NtrC protein, TyrR has intrinsic ATPase activity. Here, we report that TyrR possesses phosphatase activity. This activity is subject to inhibition by l-tyrosine and its analogues and by ATP and ATP analogues. Zinc ion (2 mM) stimulated the phosphatase activity of the TyrR protein by a factor of 57. The phosphatase-active site of TyrR was localized to a 31-kDa domain (residues 191 to 467) of the protein. However, mutational alteration of distant amino acid residues at both the N terminus and the C terminus of TyrR altered the phosphatase activity. Haemophilus influenzae TyrR (318 amino acid residues), a protein with a high degree of sequence similarity to the C terminus of the E. coli TyrR protein, exhibited a phosphatase activity similar to that of E. coliTyrR.

scholarly journals Infectivity of the Highly Transformable BBE02− lp56− Mutant of Borrelia burgdorferi, the Lyme Disease Spirochete, via Ticks

2006 ◽  
Vol 74 (6) ◽  
pp. 3678-3681 ◽  
Author(s):  
Mary B. Jacobs ◽  
Steven J. Norris ◽  
Kathrine M. Phillippi-Falkenstein ◽  
Mario T. Philipp
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Ixodes Scapularis ◽  
Shuttle Vector ◽  
Lower Number ◽  
Linear Plasmid ◽  
Wild Type ◽  
Gene Encoding ◽  
Restriction Modification ◽  
Feeding Tick

ABSTRACT Infectious Borrelia burgdorferi strains that have increased transformability with the shuttle vector pBSV2 were recently constructed by inactivating the gene encoding BBE02, a putative restriction-modification gene product expressed by the linear plasmid lp25 (Kawabata et al., Infect. Immun. 72:7147-7154, 2004). The absence of the linear plasmid lp56, which carries another putative restriction-modification gene, further enhanced transformation rates. The infectivity of these mutants was assessed previously in mice that were inoculated with needle and syringe and was found to be equivalent to that of wild-type spirochetes. Here we examined the infectivity of spirochetes to ticks after capillary inoculation of Ixodes scapularis nymphs and the subsequent spirochetal infectivity to mice via ticks by using B. burgdorferi B31 clonal isolates lacking lp56 and/or BBE02. The absence of lp56 (but not BBE02) correlated with a lower number of spirochetes in ticks after feeding on mice; this plasmid thus may play a role, albeit not an essential one, in supporting spirochetal survival in the feeding tick. Importantly, however, the absence of lp56 and BBE02 did not detectably influence infectivity to mice via ticks.

scholarly journals Camelpox virus encodes a schlafen-like protein that affects orthopoxvirus virulence

2007 ◽  
Vol 88 (6) ◽  
pp. 1667-1676 ◽  
Author(s):  
Caroline Gubser ◽  
Rory Goodbody ◽  
Andrea Ecker ◽  
Gareth Brady ◽  
Luke A. J. O'Neill ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Sequence Similarity ◽  
Small Animal ◽  
Wild Type ◽  
Recombinant Viruses ◽  
C Terminus ◽  
N Terminus ◽  
Type V

Camelpox virus (CMLV) gene 176R encodes a protein with sequence similarity to murine schlafen (m-slfn) proteins. In vivo, short and long members of the m-slfn family inhibited T-cell development, whereas in vitro, only short m-slfns caused arrest of fibroblast growth. CMLV 176 protein (v-slfn) is most closely related to short m-slfns; however, when expressed stably in mammalian cells, v-slfn did not inhibit cell growth. v-slfn is a predominantly cytoplasmic 57 kDa protein that is expressed throughout infection. Several other orthopoxviruses encode v-slfn proteins, but the v-slfn gene is fragmented in all sequenced variola virus and vaccinia virus (VACV) strains. Consistent with this, all 16 VACV strains tested do not express a v-slfn detected by polyclonal serum raised against the CMLV protein. In the absence of a small animal model to study CMLV pathogenesis, the contribution of CMLV v-slfn to orthopoxvirus virulence was studied via its expression in an attenuated strain of VACV. Recombinant viruses expressing wild-type v-slfn or v-slfn tagged at its C terminus with a haemagglutinin (HA) epitope were less virulent than control viruses. However, a virus expressing v-slfn tagged with the HA epitope at its N terminus had similar virulence to controls, implying that the N terminus has an important function. A greater recruitment of lymphocytes into infected lung tissue was observed in the presence of wild-type v-slfn but, interestingly, these cells were less activated. Thus, v-slfn is an orthopoxvirus virulence factor that affects the host immune response to infection.

scholarly journals Structural mechanisms underlying sequence-dependent variations in GAG affinities of decorin binding protein A, a Borrelia burgdorferi adhesin

2015 ◽  
Vol 467 (3) ◽  
pp. 439-451 ◽  
Author(s):  
Ashli M. Morgan ◽  
Xu Wang
Keyword(s):  
Borrelia Burgdorferi ◽  
Binding Protein ◽  
Protein A ◽  
Binding Pocket ◽  
Sulfated Polysaccharide ◽  
Solution Structures ◽  
C Terminus ◽  
Binding Epitope ◽  
Different Strains ◽  
Structural Mechanisms

Decorin-binding protein A (DBPA) is an important surface adhesin of the bacterium Borrelia burgdorferi, the causative agent of Lyme disease. DBPA facilitates the bacteria's colonization of human tissue by adhering to glycosaminoglycan (GAG), a sulfated polysaccharide. Interestingly, DBPA sequence variation among different strains of Borrelia spirochetes is high, resulting in significant differences in their GAG affinities. However, the structural mechanisms contributing to these differences are unknown. We determined the solution structures of DBPAs from strain N40 of B. burgdorferi and strain PBr of Borrelia garinii, two DBPA variants whose GAG affinities deviate significantly from strain B31, the best characterized version of DBPA. Our structures revealed that significant differences exist between PBr DBPA and B31/N40 DBPAs. In particular, the C-terminus of PBr DBPA, unlike C-termini from B31 and N40 DBPAs, is positioned away from the GAG-binding pocket and the linker between helices one and two of PBr DBPA is highly structured and retracted from the GAG-binding pocket. The repositioning of the C-terminus allowed the formation of an extra GAG-binding epitope in PBr DBPA and the retracted linker gave GAG ligands more access to the GAG-binding epitopes than other DBPAs. Characterization of GAG ligands' interactions with wild-type (WT) PBr and mutants confirmed the importance of the second major GAG-binding epitope and established the fact that the two epitopes are independent of one another and the new epitope is as important to GAG binding as the traditional epitope.

scholarly journals Gene conversions are frequent but not under positive selection in the Siglec gene families of primates

Genome ◽  
2014 ◽  
Vol 57 (6) ◽  
pp. 317-325 ◽  
Author(s):  
Mouldi Zid ◽  
Guy Drouin
Keyword(s):  
Sequence Similarity ◽  
Purifying Selection ◽  
Gene Families ◽  
Surface Proteins ◽  
Primate Species ◽  
Immunoglobulin Superfamily ◽  
Sialic Acid Binding ◽  
High Degree ◽  
Gene Conversions

Siglecs are cell surface proteins that belong to the immunoglobulin superfamily and which bind sialic acids. They are composed of two groups, the conserved Siglecs and the CD33-related Siglecs. Previous studies have reported the occurrence of gene conversions between human CD33-related Siglecs and suggested that these conversions are adaptive because they increase the diversity of these immunoglobulin-related genes. Here, we analyze the Siglec genes of five primate species and show that gene conversions are not observed between conserved Siglec genes but that they are frequent between primate CD33-related Siglecs. The gene conversions between CD33-related Siglec genes only occur between similar genes and equally frequently in sialic acid binding and nonbinding domains. Furthermore, dN/dS ratio tests show that most of the Ig-like V-type 1 and the Ig-like C2-type 1 domains of Siglec genes evolve either neutrally or under purifying selection and that gene conversions were not responsible for the positively selected regions detected in the Ig-like V-type1 domain of the human SIGLEC7 and SIGLEC9 genes. Our results suggest that the frequent gene conversions between CD33-related Siglec genes are simply a consequence of the high degree of sequence similarity of these genes and that they are not adaptive.

scholarly journals CheX Is a Phosphorylated CheY Phosphatase Essential for Borrelia burgdorferi Chemotaxis

2005 ◽  
Vol 187 (23) ◽  
pp. 7963-7969 ◽  
Author(s):  
M. A. Motaleb ◽  
Michael R. Miller ◽  
Chunhao Li ◽  
Richard G. Bakker ◽  
Stuart F. Goldstein ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Environmental Changes ◽  
Capillary Tube ◽  
Bacterial Species ◽  
Shuttle Vector ◽  
Gel Filtration ◽  
Signal Transduction Pathway ◽  
Wild Type ◽  
Mutant Cells

ABSTRACT Motility and chemotaxis are believed to be important in the pathogenesis of Lyme disease caused by the spirochete Borrelia burgdorferi. Controlling the phosphorylation state of CheY, a response regulator protein, is essential for regulating bacterial chemotaxis and motility. Rapid dephosphorylation of phosphorylated CheY (CheY-P) is crucial for cells to respond to environmental changes. CheY-P dephosphorylation is accomplished by one or more phosphatases in different species, including CheZ, CheC, CheX, FliY, and/or FliY/N. Only a cheX phosphatase homolog has been identified in the B. burgdorferi genome. However, a role for cheX in chemotaxis has not been established in any bacterial species. Inactivating B. burgdorferi cheX by inserting a flgB-kan cassette resulted in cells (cheX mutant cells) with a distinct motility phenotype. While wild-type cells ran, paused (stopped or flexed), and reversed, the cheX mutant cells continuously flexed and were not able to run or reverse. Furthermore, swarm plate and capillary tube chemotaxis assays demonstrated that cheX mutant cells were deficient in chemotaxis. Wild-type chemotaxis and motility were restored when cheX mutant cells were complemented with a shuttle vector expressing CheX. Furthermore, CheX dephosphorylated CheY3-P in vitro and eluted as a homodimer in gel filtration chromatography. These findings demonstrated that B. burgdorferi CheX is a CheY-P phosphatase that is essential for chemotaxis and motility, which is consistent with CheX being the only CheY-P phosphatase in the B. burgdorferi chemotaxis signal transduction pathway.

scholarly journals Identification of Lysine Residues in the Borrelia burgdorferi DbpA Adhesin Required for Murine Infection

2014 ◽  
Vol 82 (8) ◽  
pp. 3186-3198 ◽  
Author(s):  
Danielle E. Fortune ◽  
Yi-Pin Lin ◽  
Ranjit K. Deka ◽  
Ashley M. Groshong ◽  
Brendan P. Moore ◽  
...  
Keyword(s):  
Nmr Spectroscopy ◽  
Borrelia Burgdorferi ◽  
Cellular Localization ◽  
Dermatan Sulfate ◽  
Protein A ◽  
Structural Data ◽  
Content Type ◽  
C Terminus ◽  
Lysine Residues

ABSTRACTDecorin-binding protein A (DbpA) ofBorrelia burgdorferimediates bacterial adhesion to heparin and dermatan sulfate associated with decorin. Lysines K82, K163, and K170 of DbpA are known to be important forin vitrointeraction with decorin, and the DbpA structure, initially solved by nuclear magnetic resonance (NMR) spectroscopy, suggests these lysine residues colocalize in a pocket near the C terminus of the protein. In the current study, we solved the structure of DbpA fromB. burgdorferistrain 297 using X-ray crystallography and confirmed the existing NMR structural data.In vitrobinding experiments confirmed that recombinant DbpA proteins with mutations in K82, K163, or K170 did not bind decorin, which was due to an inability to interact with dermatan sulfate. Most importantly, we determined that thein vitrobinding defect observed upon mutation of K82, K163, or K170 in DbpA also led to a defect during infection. The infectivity ofB. burgdorferiexpressing individualdbpAlysine point mutants was assessed in mice challenged via needle inoculation. Murine infection studies showed that strains expressingdbpAwith mutations in K82, K163, and K170 were significantly attenuated and could not be cultured from any tissue. Proper expression and cellular localization of the mutated DbpA proteins were examined, and NMR spectroscopy determined that the mutant DbpA proteins were structurally similar to wild-type DbpA. Taken together, these data showed that lysines K82, K163, and K170 potentiate the binding of DbpA to dermatan sulfate and that an interaction(s) mediated by these lysines is essential forB. burgdorferimurine infection.

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