Bule, P, Alves VD, Israeli-Ruimy V, Carvalho AL, Ferreira LMA, Smith SP, Gilbert HJ, Najmudin S, Bayer EA, Fontes CMGA.
2017.
Assembly of Ruminococcus flavefaciens cellulosome revealed by structures of two cohesin-dockerin complexes, 2017. Scientific Reports. 7:759.
AbstractCellulosomes are sophisticated multi-enzymatic nanomachines produced by anaerobes to effectively deconstruct plant structural carbohydrates. Cellulosome assembly involves the binding of enzyme-borne dockerins (Doc) to repeated cohesin (Coh) modules located in a non-catalytic scaffoldin. Docs appended to cellulosomal enzymes generally present two similar Coh-binding interfaces supporting a dual-binding mode, which may confer increased positional adjustment of the different complex components. Ruminococcus flavefaciens’ cellulosome is assembled from a repertoire of 223 Doc-containing proteins classified into 6 groups. Recent studies revealed that Docs of groups 3 and 6 are recruited to the cellulosome via a single-binding mode mechanism with an adaptor scaffoldin. To investigate the extent to which the single-binding mode contributes to the assembly of R. flavefaciens cellulosome, the structures of two group 1 Docs bound to Cohs of primary (ScaA) and adaptor (ScaB) scaffoldins were solved. The data revealed that group 1 Docs display a conserved mechanism of Coh recognition involving a single-binding mode. Therefore, in contrast to all cellulosomes described to date, the assembly of R. flavefaciens cellulosome involves single but not dual-binding mode Docs. Thus, this work reveals a novel mechanism of cellulosome assembly and challenges the ubiquitous implication of the dual-binding mode in the acquisition of cellulosome flexibility.
Gomes, AS, Trovão F, Andrade Pinheiro B, Freire F, Gomes S, Oliveira C, Domingues L, Romão MJ, Saraiva L, Carvalho AL.
2018.
The Crystal Structure of the R280K Mutant of Human p53 Explains the Loss of DNA Binding. International Journal of Molecular Sciences. 19, Number 4}, ARTICLE NUMBER = {1184
AbstractThe p53 tumor suppressor is widely found to be mutated in human cancer. This protein is regarded as a molecular hub regulating different cell responses, namely cell death. Compelling data have demonstrated that the impairment of p53 activity correlates with tumor development and maintenance. For these reasons, the reactivation of p53 function is regarded as a promising strategy to halt cancer. In the present work, the recombinant mutant p53R280K DNA binding domain (DBD) was produced for the first time, and its crystal structure was determined in the absence of DNA to a resolution of 2.0 Å. The solved structure contains four molecules in the asymmetric unit, four zinc(II) ions, and 336 water molecules. The structure was compared with the wild-type p53 DBD structure, isolated and in complex with DNA. These comparisons contributed to a deeper understanding of the mutant p53R280K structure, as well as the loss of DNA binding related to halted transcriptional activity. The structural information derived may also contribute to the rational design of mutant p53 reactivating molecules with potential application in cancer treatment.
Archer, M, Huber R, Tavares P, Moura I, Moura JJG, Carrondo MA, Sieker LC, Legall J, Romao MJ.
1995.
CRYSTAL-STRUCTURE OF DESULFOREDOXIN FROM DESULFOVIBRIO-GIGAS DETERMINED AT 1.8 ANGSTROM RESOLUTION - A NOVEL NONHEME IRON PROTEIN-STRUCTURE. Journal of Molecular Biology. 251:690-702., Number 5
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Romao, MJ, Archer M, Moura I, Moura JJG, Legall J, Engh R, Schneider M, Hof P, Huber R.
1995.
CRYSTAL-STRUCTURE OF THE XANTHINE OXIDASE-RELATED ALDEHYDE OXIDOREDUCTASE FROM D-GIGAS. Science. 270:1170-1176., Number 5239
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Teixeira, S, Dias JM, Carvalho AL, Bourenkov G, Bartunik H, Almendra MJ, Moura I, Moura JJG, Romao MJ.
1999.
Crystallographic studies on a tungsten-containning formate dehydrogenase from Desulfovibrio gigas. Journal of Inorganic Biochemistry. 74:89-89., Number 1-4
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Cunha, CA, Macieira S, Dias JM, Almeida G, Goncalves LL, Costa C, Lampreia J, Huber R, Moura JJG, Moura I, Romao MJ.
2003.
Cytochrome c nitrite reductase from Desulfovibrio desulfuricans ATCC 27774 - The relevance of the two calcium sites in the structure of the catalytic subunit (NrfA). Journal of Biological Chemistry. 278:17455-17465., Number 19
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Brás, JLA, Pinheiro BA, Cameron K, Cuskin F, Viegas A, Najmudin S, Bule P, Pires VMR, Romão MJ, Bayer EA, Spencer HL, Smith S, Gilbert HJ, Alves VD, Carvalho AL, Fontes CMGA.
2016.
Diverse specificity of cellulosome attachment to the bacterial cell surface, dec. Scientific Reports. 6:38292.: The Author(s)
AbstractDuring the course of evolution, the cellulosome, one of Nature's most intricate multi-enzyme complexes, has been continuously fine-tuned to efficiently deconstruct recalcitrant carbohydrates. To facilitate the uptake of released sugars, anaerobic bacteria use highly ordered protein-protein interactions to recruit these nanomachines to the cell surface. Dockerin modules located within a non-catalytic macromolecular scaffold, whose primary role is to assemble cellulosomal enzymatic subunits, bind cohesin modules of cell envelope proteins, thereby anchoring the cellulosome onto the bacterial cell. Here we have elucidated the unique molecular mechanisms used by anaerobic bacteria for cellulosome cellular attachment. The structure and biochemical analysis of five cohesin-dockerin complexes revealed that cell surface dockerins contain two cohesin-binding interfaces, which can present different or identical specificities. In contrast to the current static model, we propose that dockerins utilize multivalent modes of cohesin recognition to recruit cellulosomes to the cell surface, a mechanism that maximises substrate access while facilitating complex assembly.
Duarte, M, Viegas A, Alves VD, Prates JAM, Ferreira LMA, Najmudin S, Cabrita EJ, Carvalho AL, Fontes CMGA, Bule P.
2021.
A dual cohesin–dockerin complex binding mode in Bacteroides cellulosolvens contributes to the size and complexity of its cellulosome. Journal of Biological Chemistry. 296:100552.
AbstractThe Cellulosome is an intricate macromolecular protein complex that centralizes the cellulolytic efforts of many anaerobic microorganisms through the promotion of enzyme synergy and protein stability. The assembly of numerous carbohydrate processing enzymes into a macromolecular multiprotein structure results from the interaction of enzyme-borne dockerin modules with repeated cohesin modules present in noncatalytic scaffold proteins, termed scaffoldins. Cohesin–dockerin (Coh-Doc) modules are typically classified into different types, depending on structural conformation and cellulosome role. Thus, type I Coh-Doc complexes are usually responsible for enzyme integration into the cellulosome, while type II Coh-Doc complexes tether the cellulosome to the bacterial wall. In contrast to other known cellulosomes, cohesin types from Bacteroides cellulosolvens, a cellulosome-producing bacterium capable of utilizing cellulose and cellobiose as carbon sources, are reversed for all scaffoldins, i.e., the type II cohesins are located on the enzyme-integrating primary scaffoldin, whereas the type I cohesins are located on the anchoring scaffoldins. It has been previously shown that type I B. cellulosolvens interactions possess a dual-binding mode that adds flexibility to scaffoldin assembly. Herein, we report the structural mechanism of enzyme recruitment into B. cellulosolvens cellulosome and the identification of the molecular determinants of its type II cohesin–dockerin interactions. The results indicate that, unlike other type II complexes, these possess a dual-binding mode of interaction, akin to type I complexes. Therefore, the plasticity of dual-binding mode interactions seems to play a pivotal role in the assembly of B. cellulosolvens cellulosome, which is consistent with its unmatched complexity and size.
Ribeiro, T, Santos-Silva T, Alves VD, Dias FMV, Luis AS, Prates JAM, Ferreira LMA, Romao MJ, Fontes CMGA.
2010.
Family 42 carbohydrate-binding modules display multiple arabinoxylan-binding interfaces presenting different ligand affinities. Biochimica Et Biophysica Acta-Proteins and Proteomics. 1804:2054-2062., Number 10
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Santos-Silva, T, Trincao J, Carvalho AL, Bonifacio C, Auchere F, Raleiras P, Moura I, Moura JJG, Romao MJ.
2006.
The first crystal structure of class III superoxide reductase from Treponema pallidum. Journal of Biological Inorganic Chemistry. 11:548-558., Number 5
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Rebelo, J, Macieira S, Dias JM, Huber R, Ascenso CS, Rusnak F, Moura JJG, Moura I, Romao MJ.
2000.
Gene sequence and crystal structure of the aldehyde oxidoreductase from Desulfovibrio desulfuricans ATCC 27774. Journal of Molecular Biology. 297:135-146., Number 1
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Bule, P, Pires VMR, Alves VD, Carvalho AL, Prates JAM, Ferreira LMA, Smith SP, Gilbert HJ, Noach I, Bayer EA, Najmudin S, Fontes CMGA.
2018.
Higher order scaffoldin assembly in Ruminococcus flavefaciens cellulosome is coordinated by a discrete cohesin-dockerin interaction, 2018. Scientific Reports. 8(1):6987.
AbstractCellulosomes are highly sophisticated molecular nanomachines that participate in the deconstruction of complex polysaccharides, notably cellulose and hemicellulose. Cellulosomal assembly is orchestrated by the interaction of enzyme-borne dockerin (Doc) modules to tandem cohesin (Coh) modules of a non-catalytic primary scaffoldin. In some cases, as exemplified by the cellulosome of the major cellulolytic ruminal bacterium Ruminococcus flavefaciens, primary scaffoldins bind to adaptor scaffoldins that further interact with the cell surface via anchoring scaffoldins, thereby increasing cellulosome complexity. Here we elucidate the structure of the unique Doc of R. flavefaciens FD-1 primary scaffoldin ScaA, bound to Coh 5 of the adaptor scaffoldin ScaB. The RfCohScaB5-DocScaA complex has an elliptical architecture similar to previously described complexes from a variety of ecological niches. ScaA Doc presents a single-binding mode, analogous to that described for the other two Coh-Doc specificities required for cellulosome assembly in R. flavefaciens. The exclusive reliance on a single-mode of Coh recognition contrasts with the majority of cellulosomes from other bacterial species described to date, where Docs contain two similar Coh-binding interfaces promoting a dual-binding mode. The discrete Coh-Doc interactions observed in ruminal cellulosomes suggest an adaptation to the exquisite properties of the rumen environment.
Fv, V, Violante S, Gomes C, Carvalho AL, Romao MJ, Gaspar MM, Cruz MEM, Soveral G, Wanders RJ, Leandro P, de Almeida TV.
2007.
The human carnitine acylcarnitine translocase (hCACT): Strategies for its heterologous expression, purification and crystallization. Journal of Inherited Metabolic Disease. 30:53-53.
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Vidinha, P, Lourenco NMT, Pinheiro C, Bras AR, Carvalho T, Santos-Silva T, Mukhopadhyay A, Romao MJ, Parola J, Dionisio M, Cabral JMS, Afonso CAM, Barreiros S.
2008.
Ion jelly: a tailor-made conducting material for smart electrochemical devices. Chemical Communications. :5842-5844., Number 44
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Almeida, MG, Macieira S, Goncalves LL, Huber R, Cunha CA, Romao MJ, Costa C, Lampreia J, Moura JJG, Moura I.
2003.
The isolation and characterization of cytochrome c nitrite reductase subunits (NrfA and NrfH) from Desulfovibrio desulfuricans ATCC 27774 - Re-evaluation of the spectroscopic data and redox properties. European Journal of Biochemistry. 270:3904-3915., Number 19
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Bras, JLA, Alves VD, Carvalho AL, Najmudin S, Prates JAM, Ferreira LMA, Bolam DN, Romao MJ, Gilbert HJ, Fontes CMGA.
2012.
Novel Clostridium thermocellum Type I Cohesin-Dockerin Complexes Reveal a Single Binding Mode. Journal of Biological Chemistry. 287:44394-44405., Number 53
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