Molecular masses and sedimentation coefficients of extracellular hemoglobin of Glossoscolex paulistus: Alkaline oligomeric dissociation

Carvalho, Francisco Adriano de Oliveira; Santiago, Patrícia S.; Borges, Júlio César; Tabak, Marcel

doi:10.1016/j.ijbiomac.2010.11.002

Cited by 22 publications

(27 citation statements)

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“…As initial input data, the MM determined by mass spectrometry [13] and the s-value determined by SV analysis [4] were used, and both parameters were allowed to float freely.…”

Section: Auc Data Analysismentioning

confidence: 99%

“…These extracellular hemoglobins are characterized by a very high molecular mass (MM), a high resistance to oxidation and a high oligomeric stability when subjected to conditions of stress such as high temperature, pH variation, and addition of chemical agents, such as urea and surfactants [3][4][5][6], as compared, for instance, with human hemoglobin. These properties make them an interesting and important system for investigation [2,7], including in biomedical applications.…”

Section: Introductionmentioning

confidence: 99%

“…This protein has an oligomeric structure, similar to the orthologous HbLt [12], composed by 144 globin chains, and 36 additional chains lacking the heme group, named linkers. Mass spectrometry and AUC studies suggest that HbGp has the same stoichiometry as HbLt, based on the Vinogradov model, which assumes that the whole protein is composed by twelve protomers, constituted by a dodecamer of globin chains and a trimer of linkers, [(abcd) 3 L 3 ] [1,4,13]. Here a, b, c and d are globin chains forming an asymmetric tetramer (abcd), composed of a disulfide bonded trimer (abc) and a monomeric subunit d. Three linker chains, L 1 , L 2 and L 3 complete the native protomer structure.…”

Section: Introductionmentioning

confidence: 99%

“…However, for cyanomet-HbGp, besides the appearance of these species, some fraction of undissociated whole protein is observed (17%), due to the fact that the cyanomet-form is more stable than oxy-HbGp, at pH 10.0 [4]. Spectroscopic studies of HbGp in the presence of urea suggested that urea induces, at low concentrations, between 1.0 and 3.0 mol/L, the oligomeric dissociation, and further increase in urea concentration promotes the denaturation of the dissociated species [6].…”

Section: Introductionmentioning

confidence: 99%

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Urea-induced unfolding of Glossoscolex paulistus hemoglobin, in oxy- and cyanomet-forms: A dissociation model

Carvalho

Santiago

et al. 2013

International Journal of Biological Macromolecules

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The urea effect on the giant extracellular hemoglobin of Glossoscolex paulistus (HbGp) stability was studied by analytical ultracentrifugation (AUC) and small angle X-ray scattering (SAXS). AUC data show that the sedimentation coefficient distributions curves c (S), at 1.0 mol/L of urea, display a single peak at 57 S, associated to the undissociated protein. The increase in urea concentration, up to 4.0 mol/L, induces the appearance of smaller species, due to oligomeric dissociation. The sedimentation coefficients and molecular masses are 9.2S and 204 kDa for the dodecamer (abcd)(3), 5.5S and 69 kDa for the tetramer (abcd), 4.1S and 52 kDa for the trimer (abc) and 2.0 S and 17 kDa for the monomer d, respectively. SAXS data show initially a decrease in the I(0) values due to the oligomeric dissociation, and then, above 4.0 mol/L of denaturant, for oxy-HbGp, and above 6.0 mol/L for cyanomet-HbGp, an increase in the maximum dimension and gyration radius is observed, due to the unfolding process. According to AUC and SAXS data the HbGp unfolding is described by two phases: the first one, at low urea concentration, below 4.0 mol/L, characterizes the oligomeric dissociation, while the second one, at higher urea concentration, is associated to the unfolding of dissociated species. Our results are complementary to a recent report based on spectroscopic observations.

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“…As initial input data, the MM determined by mass spectrometry [13] and the s-value determined by SV analysis [4] were used, and both parameters were allowed to float freely.…”

Section: Auc Data Analysismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Urea-induced unfolding of Glossoscolex paulistus hemoglobin, in oxy- and cyanomet-forms: A dissociation model

Carvalho

Santiago

et al. 2013

International Journal of Biological Macromolecules

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“…Além disso, estudos por ultracentrifugação analítica [20], e cromatografia de exclusão por tamanho [3,6] …”

Section: Fonte-unclassified

Estudo da estabilidade térmica da hemoglobina extracelular gigante de Glossoscolex paulistus (HbGp): efeitos do estado de oxidação do ferro do grupo heme, pH e presença de surfactante

Carvalho¹

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AgradecimentosAgradeço! Aos meus pais Maria Gorete Pires Carvalho e José de Ribamar Gomes Carvalho e a minha esposa Sumária Sousa e Silva pela dedicação, carinho, motivação nas horas mais difíceis, sem eles não teria forças para seguir essa jornada; Ao Professor Marcel Tabak por sua orientação e pelos laços de amizade e respeito que se formaram nesta convivência e também pela oportunidade de poder desenvolver esse trabalho;A Fundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP) pela bolsa de doutorado;Aos meus irmãos Genilson, Genilda e Gesilda pelo apoio, carinho e afeto que temos e pelas palavras amigas;Às minhas tias Eva (segunda mãe), Maria da Guia e Raimunda Teresa e os primos (as) Rita de Kássia, Karla Karolina, Raiza, Rairone, Railson, Amanda e Constância pelo apoio e carinho que sempre me motiva; Aos meus amigos de laboratório Adriano, Fernanda, Ana Eliza, Diógenes e Ezer Biazin, pelo apoio moral e pela contribuição direta e indireta na minha formação e neste trabalho. Obrigado em especial a vocês Adriano e Ezer Biazin; Aos meus amigos Orlando, Ana Célia, Adriane, Washington, Alexandre, Tairo, Adriana, Elenice, Jairo e Edvan pelas discussões bem humoradas e pela amizade; Aos professores Dr.(a) (s) Leandro R. S. Barbosa (IF-USP), Rosângela Itri (IF-USP), Patríca S. Santiago (UNESP-Registro), Carlos Ernesto Garrido Salmon (USP-Riberão Preto) e Tatiana Batista (UEL-PR) pelas discussões e colaborações durante meu curso de doutorado, que foi muito importante para minha formação; Ao Ezér Biazin, técnico do laboratório, por toda ajuda na parte instrumental e que sem nenhuma dúvida foi muito importante para o bom desenvolvimento deste trabalho e pela amizade que se construiu nestes cinco anos; A Andressa Patrícia Alves Pinto, técnica do laboratório de biofísica do IFSC por toda ajuda nos experimentos de CD que foi muito importante para o bom desenvolvimento deste trabalho; Ao Laboratório Nacional de Luz Síncrotron (LNLS), em especial ao Dr. Mateus Cardoso, coordenador da linha SAXS2, pelo auxílio na realização das medidas de SAXS;Ao pessoal da pós-graduação, Sílvia, Andréia e Gustavo, por toda a atenção, carinho e pelos bons serviços prestados;A dona Suely e seu José de Ribamar pelos conselhos e apoio nos momentos difíceis.

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Glossoscolex paulistus extracellular hemoglobin (HbGp) oligomeric dissociation upon interaction with sodium dodecyl sulfate: Isothermal titration calorimetry (ITC)

Alves

Carvalho

et al. 2014

Biopolymers

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Annelid erythrocruorins are respiratory proteins with high cooperativity and low autoxidation rates. The giant extracellular hemoglobin of the earthworm, Glossoscolex paulistus (HbGp), has a molecular mass of 3.6 MDa. In this work, isothermal titration calorimetry (ITC), together with DLS and fluorescence emission have been used to investigate the interaction of SDS with the HbGp in the oxy-form, at pH 7.0. Our ITC and DLS results show that addition of SDS induces oxy-HbGp oligomeric dissociation, while a small amount of protein aggregation is observed only by DLS. Moreover, the oligomeric dissociation process is favored at lower protein concentrations. The temperature effect does not influence significantly the interaction of SDS with the hemoglobin, due to the similarities presented by the critical aggregation concentration (cac) and critical micelle concentration (cmc') for the mixtures. The increase of oxy-HbGp concentration leads to a slight variation of the cac values for the SDS-oxy-HbGp mixture, attributed mainly to the noncooperative electrostatic binding of surfactant to protein. However, the cmc' values increase considerably, associated to a more cooperative hydrophobic binding. Complementary pyrene fluorescence emission studies show formation of pre-micellar structures of the mixture already at lower SDS concentrations. This study opens the possibility of the evaluation of the surfactant effect on the hemoglobin stability by ITC, which is made for the first time with this extracellular hemoglobin.

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Molecular masses and sedimentation coefficients of extracellular hemoglobin of Glossoscolex paulistus: Alkaline oligomeric dissociation

Cited by 22 publications

References 34 publications

Urea-induced unfolding of Glossoscolex paulistus hemoglobin, in oxy- and cyanomet-forms: A dissociation model

Urea-induced unfolding of Glossoscolex paulistus hemoglobin, in oxy- and cyanomet-forms: A dissociation model

Estudo da estabilidade térmica da hemoglobina extracelular gigante de Glossoscolex paulistus (HbGp): efeitos do estado de oxidação do ferro do grupo heme, pH e presença de surfactante

Glossoscolex paulistus extracellular hemoglobin (HbGp) oligomeric dissociation upon interaction with sodium dodecyl sulfate: Isothermal titration calorimetry (ITC)

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