title: Conformational dissection of Thermomyces lanuginosus lipase in solution.
abstract: Lipases are triacyl glycerol acyl hydrolases, which catalyze hydrolysis of esters, esterification and transesterification reactions, among others. Some of these enzymes have a large hydrophobic pocket covered by an alpha-helical mobile surface loop (the lid). Protein-protein interactions can occur through adsorption of two open lids of individual lipases. We investigated the conformation and oligomeric state of Thermomyces lanuginosus lipase (TLL) in solution by spectroscopic and mass spectrometry techniques. Information about oligomerization of this important industrial enzyme is only available for TLL crystals; therefore, we have done a throughout investigation of the conformation of this lipase in solution. SDS-PAGE and mass spectrometry analysis of size-exclusion chromatography eluted fractions indicated the presence of both monomeric and dimeric populations of TLL. The stability of the enzyme upon thermal and guanidine hydrochloride treatment was examined by circular dichroism and fluorescence emission spectroscopy. Small angle x-ray scattering and ion mobility mass spectrometry analysis revealed that TLL is found as a mixture of monomers and dimers at the assayed concentrations. Although previous x-ray diffraction data showed TLL as a dimer in the crystal (PDB: 1DT3), to our knowledge our report is the first evidencing that TLL co-exists as stable dimeric and monomeric forms in solution.
authors.affiliation: Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-902, Brazil; Instituto de Química, Universidade Federal do Rio de Janeiro, Centro de Tecnologia, Rio de Janeiro, RJ, Brazil.
authors.forename: Karen M
authors.initials: KM
authors.lastname: Gonçalves
authors.affiliation: Instituto de Física, Universidade de São Paulo, SP, Brazil.
authors.forename: Leandro R S
authors.initials: LR
authors.lastname: Barbosa
authors.affiliation: Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-902, Brazil; Laboratory for Structural Biology (DIMAV), Brazilian National Institute of Metrology, Quality and Technology-INMETRO, Duque de Caxias, RJ 25250-020, Brazil.
authors.forename: Luís Maurício T R
authors.initials: LM
authors.lastname: Lima
authors.affiliation: Departamento de Virologia, Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-590, Brazil.
authors.forename: Juliana R
authors.initials: JR
authors.lastname: Cortines
authors.affiliation: Laboratório Interdisciplinar de Pesquisas Médicas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ 21040-360, Brazil.
authors.forename: Dário E
authors.initials: DE
authors.lastname: Kalume
authors.affiliation: Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-902, Brazil.
authors.forename: Ivana C R
authors.initials: IC
authors.lastname: Leal
authors.affiliation: Instituto de Química, Universidade Federal do Rio de Janeiro, Centro de Tecnologia, Rio de Janeiro, RJ, Brazil.
authors.forename: Leandro S
authors.initials: LS
authors.lastname: Mariz e Miranda
authors.affiliation: Instituto de Química, Universidade Federal do Rio de Janeiro, Centro de Tecnologia, Rio de Janeiro, RJ, Brazil.
authors.forename: Rodrigo O M
authors.initials: RO
authors.lastname: de Souza
authors.affiliation: Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-902, Brazil. Electronic address: yraima@pharma.ufrj.br.
authors.forename: Yraima
authors.initials: Y
authors.lastname: Cordeiro
mesh_terms: D001203:Ascomycota / Q000737:chemistry / Q000201:enzymology*; D002942:Circular Dichroism; D008049:Lipase / Q000737:chemistry*; D013058:Mass Spectrometry; D008958:Models, Molecular; D055503:Protein Multimerization; D053838:Scattering, Small Angle; D013050:Spectrometry, Fluorescence; D014961:X-Ray Diffraction
chemical_list: D008049:Lipase
keywords: Circular dichroism; Fluorescence; Ion mobility mass spectrometry; Lipase; Oligomer; Small angle x-ray scattering