Cleavage of bovine brain microtubule-associated protein-2 by human immunodeficiency virus proteinase.
The high-molecular-weight dendritic cytoskeletal protein known as microtubule-associated protein (MAP)-2 displays the capacity to stimulate tubulin polymerization and to associate with microtubules. Serine proteases cleave MAP-2 into a C-terminal M(r) 28,000-35,000 microtubule-binding fragment and a larger N-terminal M(r) 240,000 projection-arm region. We now show that human immunodeficiency virus (HIV) proteinase also progressively degrades purified MAP-2 in vitro. This proteolysis reaction is characterized by transient accumulation of at least six intermediates, and most abundant of these is an M(r) 72,000 species that retains the ability to associate with taxol-stabilized microtubules. Treatment of this M(r) 72,000 species with thrombin releases the same M(r) 28,000 component as that derived from thrombin action on intact high-molecular-weight MAP-2, indicating that the viral aspartoproteinase action preferentially occurs further toward the N-terminus. The association of the M(r) 72,000 component with microtubules can be disrupted by the presence of a 21-amino acid peptide analogue of the second repeated sequence in the MAP-2 microtubule-binding region. We also studied HIV proteinase action on MAP-2 in the presence of tubulin and other MAPs that recycle with tubulin, and contrary to other published studies we found no effect of such treatment on microtubule self-assembly behavior. Cleavage of isolated MAP-2 by the HIV enzyme at high salt concentrations, followed by desalting and addition of tubulin, also resulted in microtubule assembly, albeit with slightly reduced efficiency.
Ainsztein
A M
AM
Department of Biochemistry and Molecular Biology, University of Florida College of Medicine, Gainesville 32610-0245.
Purich
D L
DL
D000818:Animals; D002417:Cattle; D010450:Endopeptidases / Q000378:metabolism*; D006678:HIV / Q000201:enzymology*; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008970:Molecular Weight; D009994:Osmolar Concentration; D010446:Peptide Fragments / Q000737:chemistry; D013917:Thrombin / Q000378:metabolism
D008869:Microtubule-Associated Proteins; D010446:Peptide Fragments; D010450:Endopeptidases; D013917:Thrombin