Molecular bases of memory, neurodegeneration and biocatalysis

Own lab page: http://carrionvazquez-lab.org

The general interest of our laboratory is to understand the inner workings of proteins, one of the great pending subjects of biology. Specifically, we are studying the molecular bases of memory, neurodegeneration and biocatalysis. For this, we concentrate in amyloids, both pathological (particularly neurotoxic proteins, involved in neurodegenerative diseases) and functional (CPEB, involved in memory consolidation by labelling active synapses), and scaffolding proteins (which coordinate enzymatic cascades). Our current research lines have a double aspect, basic and applied, and are the following:

1) Molecular bases of neurodegeneration: nanomechanics of the proteasome and its neurotoxic substrate proteins (huntingtin, β-amyloid, tau, α-synuclein, TDP-43 and prions).

1.1 Testing our working hypothesis that postulates the possibility that neurotoxic proteins have conformers with a mechanical stability higher than the traction power of the "unfoldase" of the proteasome (a AAA+ ATPase), which would make it difficult (or even impossible) its processing by this structure.

1.2 Identification of the initiating monomeric conformer of amyloidogenesis (missing link) by comparative analysis of mutants and changing the conditions of the medium.

1.3 Direct study of the dynamics of oligomer formation and its mechanoestability in real-time and with nanoscopic resolution.

1.4 QBP1 (an anti-amyloidogenic inhibitor peptide) as a prophylactic/ therapeutic agent in several neurodegenerative diseases: in vitro and in vivo studies (using a QBP1 transgenic mouse).

2) Molecular bases of memory consolidation: role of the functional prionoid CPEB3 from mammals.

2.1 Analysis of the high-resolution structure of human CPEB3 by solution NMR and effect of the post-translational modifications that regulate its function.

2.2 Study in vitro and in vivo of the life span of the toxic oligomers from human and mouse CPEB3s: Analysis of its toxicity and regulation as well as its pathways of necrosis and apoptosis.

2.3 Studies in vivo and in vitro of an hypothetical interaction of human CPEB3 with neurotoxic proteins by formation of hetero-oligomers: a mechanism for generation of cognitive dysfunction prior to cell death via sequestration of CPEB3.

2.4 Prophylaxis/therapy of Posttraumatic Stress Disorder by the use of QBP1: in vitro and in vivo studies using the QBP1 transgenic mouse. Use of memory paradigms to test whether consolidation is blocked.

3) Molecular bases of biocatalysis: nanomechanics of scaffolding proteins which assemble multi-enzimatic cascades (scaffoldins): theie mechanisms and applications.

3.1 Mechanostability as a new industrial parameter: use of mechanically reinforced designer miniscaffoldins for enzymatic improvement.

3.2 Analysis of potentially amyloidogenic/prionogenic linkers for enzyme improvement: elucidation of the molecular mechanism involved and possible applications.

3.3 Enzymatic improvement through "resurrected" (ancestral) proteins coupled to scaffolding proteins. Structural and activity studies to optimize the biocatalysis.

Most relevant methodology:

Our multidisciplinary approach includes, on one side, "classical" techniques such as genetic engineering, protein engineering, structural biology, computer simulations (molecular dynamics), traditional biochemistry, enzymology, cell culture and animal experimentation (Drosophila and mouse transgenic models). On the other hand, the most distinctive methodology that we use in our lab can be called "single-molecule biochemistry", since its goal is the study of the function of biomolecules, and particularly proteins, one molecule at a time. This approach allows us to analyze, with nanoscopic resolution, the dynamics of protein molecules in real time and under physiological conditions. In particular, the specific technique that we use is atomic force microscopy (AFM), in its two modalities: single-molecule force spectroscopy and imaging. This technique allows for nanomanipulation, analysis of nanomechanics, and "mechanical visualization" of individual protein molecules. Our long-term goal is to extend this methodology to study the nanomechanics of proteins inside cells and living organisms; to that end, we plan to develop molecular force sensors that would allow the measurement of mechanical forces in vivo for the specific proteins we are interested.