29 INTEGRANTES

GRUPLAC

DESCRIPCIÓN 

Las líneas de trabajo del GRUPO INVESTIGACIONES BIOMÉDICAS Y DE GENÉTICA HUMANA APLICADA GIBGA buscan generar investigación biomédica que cubra varios frentes importantes a nivel nacional.

Es el objetivo desarrollar estudios sobre:

  • las correlaciones de la estructura metabólica, la virulencia y la filogeográfica de cepas de Helicobacter pylori para la identificación de blancos terapéuticos.
  • Problemas concernientes a la fisiología y anatomía comparada humana.
  • La determinación de la genotoxicidad en muestras de personal en empleos que se encuentren en alto riesgo de exposición a genotóxicos y cancerígenos.
  • La creación de un Laboratorio de diagnóstico molecular del cáncer e identificación de genes que confieran susceptibilidad a los mismos.
  • Establecimiento de líneas celulares de cáncer de estomago, mama y pulmón e identificación de metabolitos inhibidores del crecimiento.

Nuestra orientación trata de potenciar la conexión entre la investigación básica, clínica, epidemiológica y computacional sobre temas pertinentes a nuestra región.

SEMILLEROS

PRODUCTOS DESTACADOS

Production and characterization of a human lysosomal recombinant iduronate‐2‐sulfatase produced in Pichia pastoris

Hunter syndrome (Mucopolysaccharidosis II, MPS II) is an X‐linked lysosomal storage disease produced by the deficiency of the lysosomal enzyme iduronate‐2‐sulfatase (IDS). Currently, MPS II patients are mainly treated with enzyme replacement therapy (ERT) using recombinant enzymes produced in mammalian cells. As an alternative, several studies have shown the production of active and therapeutic forms of lysosomal proteins in microorganisms. In this paper, we report the production and characterization of a recombinant IDS produced in the yeast Pichia pastoris (prIDS). We evaluated the effect of culture conditions and gene sequence optimization on prIDS production. The results showed that the highest production of prIDS was obtained at oxygen‐limited conditions using a codon‐optimized IDS cDNA. The purified enzyme showed a final activity of 12.45 nmol mg−1 H−1 and an apparent molecular mass of about 90 kDa. The highest stability was achieved at pH 6.0, and prIDS also showed high stability in human serum. Noteworthy, the enzyme was taken up by culture cells in a dose‐dependent manner through mannose receptors, which allowed the delivery of the enzyme to the lysosome. In summary, these results show the potential of Pichia pastoris as a host to produce an IDS intended for a MPS II ERT.

PREVALENCE OF DISTAL MEDIAL STRIATAL ARTERY IN A SAMPLE OF COLOMBIAN AND PERUVIAN POPULATIONS

Prevalence of Distal Medial Striatal Artery in a Sample of Colombian and Peruvian Populations

Mismatch between perceived family and individual chronotype and their association with sleep-wake patterns

While social zeitgebers are known to shape diurnal preference, little research has been devoted to determining the contribution of the familiar group chronotype as social zeitgeber on individual circadian rhythms and sleep-wake patterns in adult subjects. The current study aimed to examine the matching between perceived family chronotype and individual chronotype and their relationship with sleep-wake patterns on weekdays and weekends, diurnal subjective somnolence, and substance consumption. Nine hundred and forty-two Colombian adults completed the Composite Scale of Morningness, the Epworth Sleepiness Scale, and responded to a questionnaire about circadian preferences of their family nucleus. We found evidence of a mismatch between perceived family and individual chronotype, mainly for morning-type individuals (Cohen’s Kappa = −0.231; p < 0.001). This mismatch was associated with diurnal subjective somnolence (β = 0.073; p < 0.001) and specific sleep-wake patterns (p < 0.01). In addition, subjects with evening-type families showed higher caffeine and alcohol consumption (p < 0.001). To our knowledge, this is the first study to assess and report the mismatching between perceived family and individual chronotypes, and it adds to the existing body of knowledge regarding the influence of social zeitgebers on circadian rhythms. This is particularly relevant since mismatching between circadian physiology and environmental cues have been shown to lead to diverse pathologies.

Improvement in the production of the human recombinant enzyme N-acetylgalactosamine-6-sulfatase (rhGALNS) in Escherichia coli using synthetic biology approaches

Previously, we demonstrated production of an active recombinant human N-acetylgalactosamine-6-sulfatase (rhGALNS) enzyme in Escherichia coli as a potential therapeutic alternative for mucopolysaccharidosis IVA. However, most of the rhGALNS produced was present as protein aggregates. Here, several methods were investigated to improve production and activity of rhGALNS. These methods involved the use of physiologically-regulated promoters and alternatives to improve protein folding including global stress responses (osmotic shock), overexpression of native chaperones, and enhancement of cytoplasmic disulfide bond formation. Increase of rhGALNS activity was obtained when a promoter regulated under σ s was implemented. Additionally, improvements were observed when osmotic shock was applied. Noteworthy, overexpression of chaperones did not have any effect on rhGALNS activity, suggesting that the effect of osmotic shock was probably due to a general stress response and not to the action of an individual chaperone. Finally, it was observed that high concentrations of sucrose in conjunction with the physiological-regulated promoter proU mod significantly increased the rhGALNS production and activity. Together, these results describe advances in the current knowledge on the production of human recombinant enzymes in a prokaryotic system such as E. coli, and could have a significant impact on the development of enzyme replacement therapies for lysosomal storage diseases.

Identification of the iduronate-2-sulfatase proteome in wild-type mouse brain

Iduronate-2-sulfatase (IDS) is a lysosomal enzyme involved in the metabolism of the glycosaminoglycans heparan (HS) and dermatan (DS) sulfate. Mutations on IDS gene produce mucopolysaccharidosis II (MPS II), characterized by the lysosomal accumulation of HS and DS, leading to severe damage of the central nervous system (CNS) and other tissues. In this study, we used a neurochemistry and proteomic approaches to identify the brain distribution of IDS and its interacting proteins on wild-type mouse brain. IDS immunoreactivity showed a robust staining throughout the entire brain, suggesting an intracellular reactivity in nerve cells and astrocytes. By using affinity purification and mass spectrometry we identified 187 putative IDS partners-proteins, mainly hydrolases, cytoskeletal proteins, transporters, transferases, oxidoreductases, nucleic acid binding proteins, membrane traffic proteins, chaperons and enzyme modulators, among others. The interactions with some of these proteins were predicted by using bioinformatics tools and confirmed by co-immunoprecipitation analysis and Blue Native PAGE. In addition, we identified cytosolic IDS-complexes containing proteins from predicted highly connected nodes (hubs), with molecular functions including catalytic activity, redox balance, binding, transport, receptor activity and structural molecule activity. The proteins identified in this study would provide new insights about IDS physiological role into the CNS and its potential role in the brain-specific protein networks.

GALERÍA