Analysis of Amino Acid Sequence Variations and Immunoglobulin E-Binding Epitopes of German Cockroach Tropomyosin

ABSTRACT The allergenicities of tropomyosins from different organisms have been reported to vary. The cDNA encoding German cockroach tropomyosin (Bla g 7) was isolated, expressed, and characterized previously. In the present study, the amino acid sequence variations in German cockroach tropomyosin were analyzed in order to investigate its influence on allergenicity. We also undertook the identification of immunodominant peptides containing immunoglobulin E (IgE) epitopes which may facilitate the development of diagnostic and immunotherapeutic strategies based on the recombinant proteins. Two-dimensional gel electrophoresis and immunoblot analysis with mouse anti-recombinant German cockroach tropomyosin serum was performed to investigate the isoforms at the protein level. Reverse transcriptase PCR (RT-PCR) was applied to examine the sequence diversity. Eleven different variants of the deduced amino acid sequences were identified by RT-PCR. German cockroach tropomyosin has only minor sequence variations that did not seem to affect its allergenicity significantly. These results support the molecular basis underlying the cross-reactivities of arthropod tropomyosins. Recombinant fragments were also generated by PCR, and IgE-binding epitopes were assessed by enzyme-linked immunosorbent assay. Sera from seven patients revealed heterogeneous IgE-binding responses. This study demonstrates multiple IgE-binding epitope regions in a single molecule, suggesting that full-length tropomyosin should be used for the development of diagnostic and therapeutic reagents.

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Conformational and Linear B-Cell Epitopes of Asp f 2, a Major Allergen of Aspergillus fumigatus, Bind Differently to Immunoglobulin E Antibody in the Sera of Allergic Bronchopulmonary Aspergillosis Patients

Infection and Immunity ◽

10.1128/iai.67.5.2284-2291.1999 ◽

1999 ◽

Vol 67 (5) ◽

pp. 2284-2291 ◽

Cited By ~ 32

Author(s):

Banani Banerjee ◽

Paul A. Greenberger ◽

Jordan N. Fink ◽

Viswanath P. Kurup

Keyword(s):

Amino Acid ◽

Aspergillus Fumigatus ◽

B Cell ◽

Immunoglobulin E ◽

Allergic Bronchopulmonary Aspergillosis ◽

Amino Acid Sequences ◽

Epitope Region ◽

B Cell Epitopes ◽

Ige Binding ◽

Binding Regions

ABSTRACT Asp f 2 is a major Aspergillus fumigatus allergen involved in allergic bronchopulmonary aspergillosis. Knowledge of the B-cell epitopes may contribute to the understanding of immunoregulation and immunodiagnosis. To elucidate the immunoglobulin E (IgE) binding epitopes in the linear sequence of Asp f 2, we synthesized decamer peptides spanning the whole molecule of Asp f 2 on derivatized cellulose membranes and evaluated IgE binding in ABPA patient and control sera. Peptides three to five amino acids long were synthesized based on amino acid sequences within the IgE binding regions and evaluated for the specificity of epitope antibody interactions. Nine IgE binding regions were recognized in this protein of 268 amino acid residues. Of the nine epitopes, seven (ATQRRQI, RKYFG, HWR, YTTRR, DHFAD, ALEAYA, and THEGGQ) are present in the hydrophilic regions of Asp f 2. Immunologic evaluation of the three recombinant fragments, Asp f 2A encompassing the N-terminal epitope region, Asp f 2B without N- and C-terminal regions of the protein, and Asp f 2C representing C-terminal epitopes, revealed that either the N- or C-terminal region of the protein is essential for the correct folding and conformation for IgE antibody binding.

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Types A and B of Beet necrotic yellow vein virus Occur in Poland

Plant Disease ◽

10.1094/pd-90-1261b ◽

2006 ◽

Vol 90 (9) ◽

pp. 1261-1261 ◽

Cited By ~ 1

Author(s):

N. Borodynko

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Sugar Beet ◽

Amino Acid Sequence Identity ◽

Type A ◽

Yellow Vein ◽

Enzyme Linked Immunosorbent Assay ◽

Type B ◽

Rt Pcr ◽

Sequence Identity

Beet necrotic yellow vein virus (BNYVV) is the type member of the genus Benyvirus and the causal agent of rhizomania disease of sugar beet. Since 1999, BNYVV is becoming one of the most important viruses of sugar beet in Poland. BNYVV is represented by three types, frequently A and B and rarely P, however, in Poland, type A only was recorded (2). In 2005, a survey was conducted to determine the incidence of types A and B of BNYVV in Poland. Thirty samples of sugar beet plants with rhizomania-like symptoms were collected from six fields in the western, southern, and northern areas of Poland. All samples were analyzed by double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) with commercial antiserum (Bio-Rad, Hercules, CA). Infection of BNYVV was found in 26 samples. The presence of the virus in these samples was confirmed by reverse transcription-polymerase chain reaction (RT-PCR). The total RNA extracted from sugar beet was tested with specific primers designed to amplify a fragment of the RNA2 for BNYVV (1). Multiplex (m) RT-PCR with two sets of primers, rhizo AF/rhizo AR and rhizo BF/rhizo BR (2), was used to distinguish A and B types of BNYVV. Two obtained mRT-PCR products were sequenced and compared with other sequences available in GenBank. A 121-nt amplicon sequence (GenBank Accession No. DQ 228872) had 100% nucleotide and amino acid sequence identity with all BNYVV type B sequences (e.g., Stourton-GenBank Accession No. AY682695) (2). A 171-nt amplicon sequence (GenBank Accession No. DQ228871) had 100% nucleotide and amino acid sequence identity with all BNYVV type A sequences (e.g., Ravenna-GenBank Accession No. AY 654282) (2). Type A was detected in 23 BNYVV-infected sugar beet plants from five fields located in western and southern Poland while type B was found in three samples from one field in northern Poland. To my knowledge, this is the first report of the natural occurrence of BNYVV type B in Poland. References: (1) A. Maunier et al. Appl. Environ. Microbiol. 69:2356, 2003. (2) C. Ratti et al. J. Virol. Methods 124:41, 2005.

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Variants of Triticum mosaic virus Isolated From Wheat in Colorado Show Divergent Biological Behavior

Plant Disease ◽

10.1094/pdis-10-12-0925-re ◽

2013 ◽

Vol 97 (7) ◽

pp. 903-911 ◽

Cited By ~ 3

Author(s):

Dallas L. Seifers ◽

Satyanarayana Tatineni ◽

Roy French

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Amino Acid Sequence ◽

Mosaic Virus ◽

Dry Weight ◽

Amino Acid Sequences ◽

Biological Behavior ◽

Enzyme Linked Immunosorbent Assay ◽

Temperature Sensitive ◽

Coat Proteins

Triticum mosaic virus (TriMV) is a recently discovered virus infecting wheat. We compared Colorado isolates C10-492 and C11-775 with the 06-123 isolate. Comparisons were made using enzyme-linked immunosorbent assay (ELISA), infectivity assay, host range, dry weight (DW), inoculation of ‘Mace’ wheat with temperature-sensitive resistance to Wheat streak mosaic virus, and the deduced amino acid sequence of the coat proteins (CP) and P1 proteins. Both C10-492 and C11-775 infected ‘Gallatin’ barley and, when compared with 06-123, had significantly reduced ELISA values and virus titers in wheat. Both Colorado isolates caused symptomless infections in Mace, whereas 06-123 caused mosaic symptoms. The amino acid sequences of the CP differed by two and one amino acids for C10-492 and C11-775, respectively, compared with 06-123. The sequence of C10-492 differed from C11-775 by one amino acid. The P1 amino acid sequence of C10-492 and C11-775 differed from 06-123 by three and one amino acids, respectively. The C10-492 and C11-775 isolates reduced DW significantly in ‘Karl 92’ but significantly less than 06-123. In ‘2317’ wheat, the Colorado isolates did not consistently cause significant reduction in DW, while 06-123 did. The data collectively indicate that C10-492 and C11-775 are isolates of TriMV showing biological behavior diverse from that of 06-123.

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Sequence variation of the HVR1 region of Hepatitis C virus in response to interferon-α and ribavirin treatment

The Journal of Infection in Developing Countries ◽

10.3855/jidc.1806 ◽

2011 ◽

Vol 5 (05) ◽

pp. 370-376 ◽

Cited By ~ 5

Author(s):

Ahmed Ali Al-Qahtani ◽

Salvatore Rubino ◽

Mohammed N. Al-Ahdal

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Amino Acid ◽

Amino Acid Sequence ◽

Sequence Variation ◽

Amino Acid Sequences ◽

Interferon Α ◽

Genotype 4 ◽

Treatment Protocols ◽

Sequence Variations

Introduction: Hepatitis C virus (HCV), which is a major cause of liver diseases worldwide, undergoes genetic variation during the course of infection. The aim of this study was to examine sequence variations within the HVR1 region of HCV genotype 4 in infected Saudi patients treated with a combination therapy of interferon-α and ribavirin. Methodology: cDNA of the HVR1 region of HVC-4 from one responder and one non-responder patients was generated, cloned and sequenced. Ten clones were randomly selected and analyzed for changes in nucleotide and amino acid sequences before the start of treatment, and subsequently three and six months after the start of the therapy course. Results: Based on nucleotide and amino acid sequence variations, the HVR1 region is highly sequence variable. In both the responder and the non-responder patients, amino acid sequence variations were observed and a clear distinction between patients was evident. The amino acid changes after the treatment course were different in the responder compared to the non-responder subject. Five amino acids (residues 364 to 367, 381 and 409) were unique in the non-responder patient. Conclusion: Considerable amino acid variations were observed in the HVR1 region in both responder and non-responder patients. These findings could have implications for the development of an HCV vaccine as well as treatment protocols for HCV infections.

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Roles of the C-Terminal Amino Acids of Non-Hexameric Helicases: Insights from Escherichia coli UvrD

International Journal of Molecular Sciences ◽

10.3390/ijms22031018 ◽

2021 ◽

Vol 22 (3) ◽

pp. 1018

Author(s):

Hiroaki Yokota

Keyword(s):

Escherichia Coli ◽

Amino Acids ◽

Amino Acid ◽

Single Molecule ◽

Underlying Mechanism ◽

Amino Acid Sequences ◽

E Coli ◽

X Ray Crystallography ◽

Terminal Amino

Helicases are nucleic acid-unwinding enzymes that are involved in the maintenance of genome integrity. Several parts of the amino acid sequences of helicases are very similar, and these quite well-conserved amino acid sequences are termed “helicase motifs”. Previous studies by X-ray crystallography and single-molecule measurements have suggested a common underlying mechanism for their function. These studies indicate the role of the helicase motifs in unwinding nucleic acids. In contrast, the sequence and length of the C-terminal amino acids of helicases are highly variable. In this paper, I review past and recent studies that proposed helicase mechanisms and studies that investigated the roles of the C-terminal amino acids on helicase and dimerization activities, primarily on the non-hexermeric Escherichia coli (E. coli) UvrD helicase. Then, I center on my recent study of single-molecule direct visualization of a UvrD mutant lacking the C-terminal 40 amino acids (UvrDΔ40C) used in studies proposing the monomer helicase model. The study demonstrated that multiple UvrDΔ40C molecules jointly participated in DNA unwinding, presumably by forming an oligomer. Thus, the single-molecule observation addressed how the C-terminal amino acids affect the number of helicases bound to DNA, oligomerization, and unwinding activity, which can be applied to other helicases.

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Partial Amino Acid Sequence of a Cellulase-Like Component with IgE-Binding Properties from Stachybotrys chartarum

International Archives of Allergy and Immunology ◽

10.1159/000076439 ◽

2004 ◽

Vol 133 (2) ◽

pp. 136-144 ◽

Cited By ~ 5

Author(s):

Marja Kärkkäinen ◽

Päivi Raunio ◽

Jaakko Rautiainen ◽

Seppo Auriola ◽

Kaj Hinke ◽

...

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Stachybotrys Chartarum ◽

Partial Amino Acid Sequence ◽

Binding Properties ◽

Ige Binding

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The amino acid sequence of cytochromes c-551 from three species of Pseudomonas

Biochemical Journal ◽

10.1042/bj1310485 ◽

1973 ◽

Vol 131 (3) ◽

pp. 485-498 ◽

Cited By ~ 113

Author(s):

R. P. Ambler ◽

Margaret Wynn

Keyword(s):

Amino Acids ◽

Pseudomonas Aeruginosa ◽

Amino Acid ◽

Cytochrome C ◽

Amino Acid Sequence ◽

Amino Acid Sequences ◽

Mitochondrial Cytochrome ◽

Cytochromes C ◽

Lending Library ◽

Single Peptide

The amino acid sequences of the cytochromes c-551 from three species of Pseudomonas have been determined. Each resembles the protein from Pseudomonas strain P6009 (now known to be Pseudomonas aeruginosa, not Pseudomonas fluorescens) in containing 82 amino acids in a single peptide chain, with a haem group covalently attached to cysteine residues 12 and 15. In all four sequences 43 residues are identical. Although by bacteriological criteria the organisms are closely related, the differences between pairs of sequences range from 22% to 39%. These values should be compared with the differences in the sequence of mitochondrial cytochrome c between mammals and amphibians (about 18%) or between mammals and insects (about 33%). Detailed evidence for the amino acid sequences of the proteins has been deposited as Supplementary Publication SUP 50015 at the National Lending Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1973), 131, 5.

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Factor D̄ of the alternative pathway of human complement. Purification, alignment and N-terminal amino acid sequences of the major cyanogen bromide fragments, and localization of the serine residue at the active site

Biochemical Journal ◽

10.1042/bj1870863 ◽

1980 ◽

Vol 187 (3) ◽

pp. 863-874 ◽

Cited By ~ 34

Author(s):

D M Johnson ◽

J Gagnon ◽

K B Reid

Keyword(s):

Amino Acid ◽

Sequence Analysis ◽

Amino Acid Sequence ◽

Alternative Pathway ◽

Amino Acid Sequences ◽

Serine Residue ◽

Amino Acid Sequence Analysis ◽

Terminal Amino Acid ◽

Terminal Amino ◽

Terminal Amino Acid Sequence

The serine esterase factor D of the complement system was purified from outdated human plasma with a yield of 20% of the initial haemolytic activity found in serum. This represented an approx. 60 000-fold purification. The final product was homogeneous as judged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis (with an apparent mol.wt. of 24 000), its migration as a single component in a variety of fractionation procedures based on size and charge, and its N-terminal amino-acid-sequence analysis. The N-terminal amino acid sequence of the first 36 residues of the intact molecule was found to be homologous with the N-terminal amino acid sequences of the catalytic chains of other serine esterases. Factor D showed an especially strong homology (greater than 60% identity) with rat ‘group-specific protease’ [Woodbury, Katunuma, Kobayashi, Titani, & Neurath (1978) Biochemistry 17, 811-819] over the first 16 amino acid residues. This similarity is of interest since it is considered that both enzymes may be synthesized in their active, rather than zymogen, forms. The three major CNBr fragments of factor D, which had apparent mol.wts. of 15 800, 6600 and 1700, were purified and then aligned by N-terminal amino acid sequence analysis and amino acid analysis. By using factor D labelled with di-[1,3-14C]isopropylphosphofluoridate it was shown that the CNBr fragment of apparent mol.wt. 6600, which is located in the C-terminal region of factor D, contained the active serine residue. The amino acid sequence around this residue was determined.

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Die primäre Proteinstruktur von Stämmen des Tabakmosaikvirus

Zeitschrift für Naturforschung B ◽

10.1515/znb-1963-1207 ◽

1963 ◽

Vol 18 (12) ◽

pp. 1032-1049 ◽

Cited By ~ 13

Author(s):

B. Wittmann-Liebold ◽

H. G. Wittmann

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Amino Acid Sequence ◽

Tobacco Mosaic Virus ◽

Mosaic Virus ◽

Amino Acid Sequences ◽

Tryptic Peptides

The amino acid sequence of dahlemense, a naturally occuring strain of tobacco mosaic virus, has been determined and compared with that of the strain vulgare (Fig. 7). In this communication the experimental details are given for the elucidation of the amino acid sequences within two tryptic peptides with 65 amino acids.

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First Report of Tomato spotted wilt virus in Blackberry Lily in North America

Plant Disease ◽

10.1094/pdis.2003.87.1.102c ◽

2003 ◽

Vol 87 (1) ◽

pp. 102-102 ◽

Cited By ~ 3

Author(s):

S. Adkins ◽

L. Breman ◽

C. A. Baker ◽

S. Wilson

Keyword(s):

North America ◽

Tomato Spotted Wilt Virus ◽

Amino Acid Sequences ◽

South Florida ◽

N Gene ◽

Enzyme Linked Immunosorbent Assay ◽

Rt Pcr ◽

First Report ◽

Tomato Spotted Wilt ◽

Wilt Virus

Blackberry lily (Belamcanda chinensis (L.) DC.) is an herbaceous perennial in the Iridaceae characterized by purple-spotted orange flowers followed by persistent clusters of black fruit. In July 2002, virus-like symptoms including chlorotic ringspots and ring patterns were observed on blackberry lily leaves on 2 of 10 plants in a south Florida ornamental demonstration garden. Inclusion body morphology suggested the presence of a Tospovirus. Tomato spotted wilt virus (TSWV) was specifically identified by serological testing using enzyme-linked immunosorbent assay (Agdia, Elkhart, IN). Sequence analysis of a nucleocapsid (N) protein gene fragment amplified by reverse transcription-polymerase chain reaction (RT-PCR) with primers TSWV723 and TSWV722 (1) from total RNA confirmed the diagnosis. Nucleotide and deduced amino acid sequences of a 579 base pair region of the RT-PCR product were 95 to 99% and 95 to 100% identical, respectively, to TSWV N-gene sequences in GenBank. Since these 2-year-old plants were grown on-site from seed, they were likely inoculated by thrips from a nearby source. Together with a previous observation of TSWV in north Florida nursery stock (L. Breman, unpublished), this represents, to our knowledge, the first report of TSWV infection of blackberry lily in North America although TSWV was observed in plants of this species in Japan 25 years ago (2). References: (1) S. Adkins, and E. N. Rosskopf. Plant Dis. 86:1310, 2002. (2) T. Yamamoto and K.-I. Ohata. Bull. Shikoku Agric. Exp. Stn. 30:39, 1977.

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