Preliminary studies of the 2D crystallization of Omp1 of Serratia marcescens: observation by atomic force microscopy in native membranes environment and reconstituted in proteolipid sheets |
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Authors: | Ruiz Neus Merino Sandra Viñas Miquel Domènech Oscar Montero M Teresa Hernández-Borrell Jordi |
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Institution: | Laboratori de Microbiologia, Campus de Bellvitge, Universitat de Barcelona E-08907-L'Hospitalet de Llobregat, Spain. |
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Abstract: | In this work the porin Omp1 of Serratia marcescens was expressed in a porin deficient mutant (Escherichia coli UH302) and its functionality studied following the accumulation of ciprofloxacin in bacteria. The protein was extracted, purified and reconstituted in proteoliposomes of different composition (lipopolysaccharide (LPS), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and, 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)). Maximum extraction of the detergent was achieved applying different steps of dialysis and centrifugation. Proteolipid sheets with different composition were spread onto mica and observed by atomic force microscopy. Two-dimensional crystal of Omp1 was not observed in any case due to low resolution achieved. Judging from the images features POPC is the most suitable phospholipid to enhance 2D lattice formation for Omp1. |
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