Hybrid enterotoxin LTA::STa proteins and their protection from degradation by in vivo association with B-subunits of Escherichia coli heat-labile enterotoxin |
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| Authors: | J Sanchez T R Hirst B E Uhlin |
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| Affiliation: | 1. Grupo de Biopolímeros y Nanoestructuras, Facultad de Ciencias Químicas, Universidad Autónoma de San Luis Potosí, Av. Dr. Manuel Nava 6, San Luis Potosí 78210, Mexico;2. Sección de Biotecnología, Centro de Investigación en Ciencias de la Salud y Biomedicina, Universidad Autónoma de San Luis Potosí, Av. Sierra Leona 550, Lomas 2a. sección, San Luis Potosí 78210, Mexico;3. Laboratorio de Ciencias Básicas, Universidad Autónoma de San Luis Potosí, Facultad de Estomatología, Av. Dr. Manuel Nava No. 2, San Luis Potosí 78290, Mexico;1. State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China;2. Laboratory of Marine Ecosystem and Biogeochemistry, Second Institute of Oceanography, State Oceanic Administration, Hangzhou, China;1. Division of Cardiovascular Medicine, Department of Internal Medicine, Abboud Cardiovascular Research Center, University of Iowa Carver College of Medicine, Iowa City, IA 52242, USA;2. Shanghai First People''s Hospital, Shanghai Jiaotong University, Shanghai 200080, China;3. Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing 210009, China;4. Department of Veterans Affairs Medical Center, Iowa City, IA 52242, USA;5. Department of Pathophysiology, School of Medicine, Shenzhen University, Shenzhen 518060, China;6. Department of Surgery, Ohio State University Medical Center, Columbus, OH 43212, USA |
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| Abstract: | Chimeric proteins exhibiting antigenic determinants of the heat-labile enterotoxin (LT) and heat-stable (STa) enterotoxins on the same molecule may provide a means to obtain immunoprophylactic and diagnostic reagents for Escherichia coli-caused diarrhea. We recently showed that fusion of two different lengths of the STa gene to the C end of the A-subunit of LT (LTA) results in LTA::STa fusion proteins as monitored by GM1-ELISA [Sanchez et al.: FEBS Lett. 208 (1986) 194-198]. Here we determine the approximate molecular size of the LTA::STa fusion proteins and provide further evidence of their hybrid nature by immunoblot analysis. Using this technique we also demonstrate that to obtain detectable amounts of these recombinant proteins it is essential to coexpress them with the respective B-subunit of LT (LTB). We propose that this dependence on coexpression reflects the association between the LTA::STa hybrids and LTB subunits. The resulting LTA::STa/LTB complexes were found in the E. coli periplasm. This indicated that the exported hybrids, once associated with LTB, were stabilized and formed molecules that behaved essentially as native LT. The protective effect exerted by the B-subunit might conceivably be extended to other LTA-derived hybrid proteins, thus allowing the fusion of other foreign peptides to LTA and their subsequent recovery in the same fashion. |
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