Carvalho, AL, A. Goyal, JAM Prates, DN Bolam, HJ Gilbert, VMR Pires, LMA Ferreira, A. Planas, MJ Romao, and CMGA Fontes. "
The family 11 carbohydrate-binding module of Clostridium thermocellum Lic26A-Cel5E accommodates beta-1,4- and beta-1,3-1,4-mixed linked glucans at a single binding site."
Journal of Biological Chemistry. 279 (2004): 34785-34793.
Abstractn/a
dos Santos, Raquel, Inês Iria, Ana M. Manuel, Ana P. Leandro, Catarina A. C. Madeira, Joao Goncalves, Ana Luísa Carvalho, and Ana Cecília Roque. "
Magnetic Precipitation: A New Platform for Protein Purification."
Biotechnology JournalBiotechnology Journal. n/a.n/a (2020): 2000151.
AbstractOne of the trends in downstream processing comprises the use of ?anything-but-chromatography? methods to overcome the current downfalls of standard packed-bed chromatography. Precipitation and magnetic separation are two techniques already proven to accomplish protein purification from complex media, yet never used in synergy. With the aim to capture antibodies directly from crude extracts, a new approach combining precipitation and magnetic separation was developed and named as affinity magnetic precipitation. A precipitation screening, based on the Hofmeister series, and a commercial precipitation kit were tested with affinity magnetic particles to assess the best condition for antibody capture from human serum plasma and clarified cell supernatant. The best conditions were obtained when using PEG3350 as precipitant at 4°C for 1h, reaching 80% purity and 50% recovery of polyclonal antibodies from plasma, and 99% purity with 97% recovery yield of anti-TNFα mAb from cell supernatants. These results show that the synergetic use of precipitation and magnetic separation can represent an alternative for the efficient capture of antibodies. This article is protected by copyright. All rights reserved
Santarsia, Sabrina, Ana Sofia Grosso, Filipa Trovão, Jesús Jiménez-Barbero, Ana Luísa Carvalho, Cristina Nativi, and Filipa Marcelo. "
Molecular recognition of a Thomsen-Friedenreich antigen mimetic targeting human galectin-3."
ChemMedChem. Aug 9. doi: 10.1002/cmdc.201800525. [Epub ahead of print] (2018).
AbstractOverexpression of the Thomsen-Friedenreich (TF) antigen in cell membrane proteins occurs in 90% of adenocarcinomas. Additionally, the binding of the TF-antigen to human galectin-3 (Gal-3), also frequently overexpressed in malignancy, promotes cancer progression and metastasis. In this context, structures that interfere with this specific interaction display the potential to prevent cancer metastasis. Herein, a multidisciplinary approach, combining the optimized synthesis of a TF-antigen mimetic with NMR, X-ray crystallography methods and isothermal titration calorimetry assays has been employed to unravel the molecular structural details that govern the Gal-3/TF-mimetic interaction. The TF-mimetic presents a binding affinity for Gal-3 similar to the TF-natural antigen and retains the binding epitope and the bioactive conformation observed for the native antigen. Furthermore, from a thermodynamic perspective a decrease in the enthalpic contribution was observed for the Gal-3/TF-mimetic complex, however this behaviour is compensated by a favourable entropy gain. From a structural perspective, these results establish our TF-mimetic as a scaffold to design multivalent solutions to potentially interfere with Gal-3 aberrant interactions and likely be used to hamper Gal-3-mediated cancer cells adhesion and metastasis.
Gomes, Ana Sara, Helena Ramos, Sara Gomes, Joana B. Loureiro, Joana Soares, Valentina Barcherini, Paola Monti, Gilberto Fronza, Carla Oliveira, Lucília Domingues, Margarida Bastos, Daniel F. A. R. Dourado, Ana Luísa Carvalho, Maria João Romão, Benedita Pinheiro, Filipa Marcelo, Alexandra Carvalho, Maria M. M. Santos, and Lucília Saraiva. "
SLMP53-1 interacts with wild-type and mutant p53 DNA-binding domain and reactivates multiple hotspot mutations." 1864.1 (2020): 129440.
AbstractBackgroundHalf of human cancers harbour TP53 mutations that render p53 inactive as a tumor suppressor. As such, reactivation of mutant (mut)p53 through restoration of wild-type (wt)-like function represents one of the most promising therapeutic strategies in cancer treatment. Recently, we have reported the (S)-tryptophanol-derived oxazoloisoindolinone SLMP53-1 as a new reactivator of wt and mutp53 R280K with in vitro and in vivo p53-dependent antitumor activity. The present work aimed a mechanistic elucidation of mutp53 reactivation by SLMP53-1.
Methods and results
By cellular thermal shift assay (CETSA), it is shown that SLMP53-1 induces wt and mutp53 R280K thermal stabilization, which is indicative of intermolecular interactions with these proteins. Accordingly, in silico studies of wt and mutp53 R280K DNA-binding domain with SLMP53-1 unveiled that the compound binds at the interface of the p53 homodimer with the DNA minor groove. Additionally, using yeast and p53-null tumor cells ectopically expressing distinct highly prevalent mutp53, the ability of SLMP53-1 to reactivate multiple mutp53 is evidenced.
Conclusions
SLMP53-1 is a p53-activating agent with the ability to directly target wt and a set of hotspot mutp53.
General Significance
This work reinforces the encouraging application of SLMP53-1 in the personalized treatment of cancer patients harboring distinct p53 status.