| Cockroach allergen Bla g 2: an unusual aspartic proteinase. | |
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MedLine Citation:
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PMID: 15990787 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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BACKGROUND: Enzymatic activity of mite, fungal, and bee venom allergens is thought to potentiate their allergenicity. Bla g 2 is a potent cockroach allergen, but despite sharing sequence homology with aspartic proteinases, it contains critical amino acid substitutions that impair proteolytic activity. The biologic function of Bla g 2 remains unclear. OBJECTIVE: We sought to investigate the effects of specific amino acid substitutions on enzymatic activity, and the peptide-binding capability of Bla g 2. METHODS: Site-directed mutagenesis was used to produce a recombinant Bla g 2 mutant (Mut) with corrected canonical triads and a flap region. Another mutant (MutF - ) was expressed after additional mutations in the flap region of Mut. Bla g 2 wild-type (Wt), Mut, and MutF - were assayed for aspartic proteinase activity, and Bla g 2 Wt was tested for pepstatin binding. RESULTS: Recombinant Bla g 2 Wt and Mut did not show enzymatic activity in a milk-clotting and hemoglobin assay. By using a modified hemoglobin assay, residual activity inhibited by pepstatin was detected for MutF - and Wt at 20 microg/mL, whereas pepsin was active at a 1000-fold lower concentration. Most of Bla g 2 binding to pepstatin-agarose was nonspecific. CONCLUSION: Residual proteolytic activity was found for Bla g 2 at concentrations of approximately 4 mM. This weak activity suggests that proteolysis is not the primary function of this allergen and that it is unlikely to contribute to the allergenicity of Bla g 2. Bla g 2 has a cleft that might specifically bind ligands other than pepstatin. |
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Authors:
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Sabina Wünschmann; Alla Gustchina; Martin D Chapman; Anna Pomés |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: The Journal of allergy and clinical immunology Volume: 116 ISSN: 0091-6749 ISO Abbreviation: J. Allergy Clin. Immunol. Publication Date: 2005 Jul |
Date Detail:
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Created Date: 2005-07-01 Completed Date: 2005-08-09 Revised Date: 2009-11-19 |
Medline Journal Info:
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Nlm Unique ID: 1275002 Medline TA: J Allergy Clin Immunol Country: United States |
Other Details:
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Languages: eng Pagination: 140-5 Citation Subset: AIM; IM |
Affiliation:
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INDOOR Biotechnologies, Inc, Charlottesville, VA 22903, USA. sabina@inbio.com |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Amino Acid Sequence Animals Aspartic Acid Endopeptidases / genetics, immunology, metabolism* Cockroaches / immunology Enzyme Activation / immunology Molecular Sequence Data Mutagenesis, Site-Directed Pepstatins / metabolism Recombinant Proteins / genetics, immunology, metabolism* |
| Chemical | |
Reg. No./Substance:
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0/Pepstatins; 0/Recombinant Proteins; EC 3.4.23.-/Aspartic Acid Endopeptidases; EC 3.4.23.-/allergen Bla g 2 |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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