RESUMEN
The global crisis provoked by the SARS-CoV-2 pandemic, and the economic and social consequences associated to the essential policies applied to contain it, necessitates the expedited development of therapeutic solutions. It is a priority to produce data both rapidly and accurately in order to identify current therapies that can be repurposed to offer protection from SARS-CoV-2 infection. As healthcare workers are both at high risk for infection and able to be readily diagnosed, they offer a potential wealth of data to be analyzed. A systematic data analysis of exposure and infection rates among healthcare workers could yield patterns identifying common protective factors, such as medications with prophylactic potential against SARS-CoV-2, that can be fast-tracked into available therapies. With results suggesting their activity against other coronaviruses and their widespread adoption, Antiretroviral cocktails could be a promising initial target for such large-scale data analysis approach.
Asunto(s)
Betacoronavirus/efectos de los fármacos , Infecciones por Coronavirus/prevención & control , Infecciones por Coronavirus/transmisión , Análisis de Datos , Personal de Salud/estadística & datos numéricos , Pandemias/prevención & control , Neumonía Viral/prevención & control , Neumonía Viral/transmisión , Profilaxis Posexposición/métodos , Profilaxis Pre-Exposición/métodos , COVID-19 , Infecciones por Coronavirus/diagnóstico , Infecciones por Coronavirus/terapia , Humanos , Neumonía Viral/diagnóstico , Neumonía Viral/terapia , SARS-CoV-2RESUMEN
Enhancement, control, and tuning of hydrolytic activity and specificity of lipases are major goals for the industry. Thermoalkaliphilic lipases from the I.5 family, with their native advantages such as high thermostability and tolerance to alkaline pHs, are a target for biotechnological applications. Although several strategies have been applied to increase lipases activity, the enhancement through protein engineering without compromising other capabilities is still elusive. Lipases from the I.5 family suffer a unique and delicate double lid restructuration to transition from a closed and inactive state to their open and enzymatically active conformation. In order to increase the activity of the wild type Geobacillus thermocatenulatus lipase 2 (BTL2) we rationally designed, based on its tridimensional structure, a mutant (ccBTL2) capable of forming a disulfide bond to lock the open state. ccBTL2 was generated replacing A191 and F206 to cysteine residues while both wild type C64 and C295 were mutated to serine. A covalently immobilized ccBTL2 showed a 3.5-fold increment in esterase activity with 0.1% Triton X-100 (2336 IU mg-1) and up to 6.0-fold higher with 0.01% CTAB (778 IU mg-1), both in the presence of oxidizing sulfhydryl agents, when compared to BTL2. The remarkable and industrially desired features of BTL2 such as optimal alkaliphilic pH and high thermal stability were not affected. The designed disulfide bond also conferred reversibility to the enhancement, as the increment on activity observed for ccBTL2 was controlled by redox pretreatments. MD simulations suggested that the most stable conformation for ccBTL2 (with the disulfide bond formed) was, as we predicted, similar to the open and active conformation of this lipase.
Asunto(s)
Proteínas Bacterianas/química , Dominio Catalítico , Cisteína/genética , Geobacillus/enzimología , Lipasa/química , Sustitución de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Cisteína/química , Disulfuros/química , Estabilidad de Enzimas , Enzimas Inmovilizadas/química , Geobacillus/genética , Lipasa/genética , Lipasa/metabolismo , Simulación de Dinámica MolecularRESUMEN
Enolase is a glycolytic and gluconeogenic enzyme also found on the surface of several eukaryotic and prokaryotic cells where it acts as plasminogen binding protein. Leishmania mexicana, one of the causative agents of Leishmaniasis, binds plasminogen and, in this parasite, enolase has been previously found associated with the external face of the plasma membrane. In this work, we show that the purified recombinant enolase has plasminogen binding activity indicating that, at the surface of the parasite, the protein may function as one of the plasminogen receptors. An internal motif (249)AYDAERKMY(257), similar to the nine amino-acid internal plasminogen-binding motif in Streptococcus pneumoniae enolase, is responsible for plasminogen interaction with the parasite enolase. Anti-enolase antibodies inhibited up to 60% of plasminogen binding on live parasites indicating that enolase act as a plasminogen receptor on the parasite. The fact that enolase acts as a possible plasminogen receptor in vivo makes this protein a promising target for therapy.