Monday, April 28, 2025
4/28/2025
Maturation of adenovirus via a new type of biochemistry - DESCRIPTION (provided by applicant): The adenovirus proteinase (AVP) is activated inside nascent virus particles; it then cleaves multiple copies of six, different virion precursor protein used in the assembly of the virion to render the virus particle infectious. We observed a conundrum in that all these proteins are sequence-independent DNA binding proteins. Given the high concentration of DNA in the virion, these proteins are essentially irreversibly bound to the viral DNA. But then, how do these proteins interact to form productive collisions, i.e. collisions that lead to binding, in the absence of three-dimensional diffusion? We solved this conundrum by showing that these biochemical reactions take place on the one-dimensional contour of the viral DNA by these proteins sliding along the DNA via one-dimensional diffusion, an unprecedented observation for protein:protein interactions. Here we ask by what biophysical mechanisms do these proteins slide on DNA? In Specific Aim 1, we present preliminary evidence that an 11-amino acid peptide (pVIc) from the C-terminus of pVI, the precursor to adenovirus protein VI, can slide by itself on DNA. We propose to characterize the parameters that enable sliding. That information will then be used in Specific Aim 2 where we present preliminary evidence that pVIc is a "molecular sled" capable not only of sliding by itself but also sliding heterologous cargoes attached to it. We go on to propose that we shall show by direct visualization, using two color total internal reflection fluorescence microscopy, how AVP is activated by substrate sliding and how active AVP-pVIc complexes process by enzyme sliding the virion precursor proteins by sliding. We shall then extend our results to see if similar protein:protein interactions occur via one-dimensional diffusion in the nucleus of mammalian cells. Preliminary evidence described in Specific Aim 1 indicated that the minimum sequence in pVIc that can slide is KRRR which is a functional nuclear location signal (NLS). Thus, presumably pVIc binds to and slides on DNA via its NLS. Many nuclear proteins have an NLS that must be on the surface of the proteins to be recognized by the nuclear import machinery. Therefore, those proteins would be expected to bind to and slide along DNA interacting on the one-dimensional contour of chromosomal DNA. This new form of biochemistry, one-dimensional biochemistry, has some unique properties. For example, if an enzyme and its substrate are optimally co-oriented by being bound to DNA via an NLS, the fraction of productive enzyme-substrate collisions mediated by sliding may be increased many orders of magnitude, possibly up to 1.0, relative to in the absence of DNA. This work presents a new paradigm for how proteinases and their substrates can interact, a new paradigm for virion maturation, a new vehicle in molecular biology, the "molecular sled," and even a new form of biochemistry that may be applicable to all bimolecular interactions that take place in the nucleus of mammalian cells.
HHS’ Tracking Accountability in Government Grants System (TAGGS) website provides access to detailed descriptions of grants, loans, aggregated direct payments and other types of financial assistance awarded by the HHS Operating Divisions (OPDIVs) and the Office of the Secretary Staff Divisions (STAFFDIVs).