Towards tricking a pathogen's protease into fighting infection: the 3D structure of a stable circularly permuted onconase variant cleavedby HIV-1 protease.
Onconase® is a highly cytotoxic amphibian homolog of Ribonuclease A. Here, we describe the construction of circularly permuted Onconase® variants by connecting the N- and C-termini of this enzyme with amino acid residues that are recognized and cleaved by the human immunodeficiency virus protease. Uncleaved circularly permuted Onconase® variants are unusually stable, non-cytotoxic and can internalize in human T-lymphocyte Jurkat cells. The structure, stability and dynamics of an intact and a cleaved circularly permuted Onconase® variant were determined by Nuclear Magnetic Resonance spectroscopy and provide valuable insight into the changes in catalytic efficiency caused by the cleavage. The understanding of the structural environment and the dynamics of the activation process represents a first step toward the development of more effective drugs for the treatment of diseases related to pathogens expressing a specific protease. By taking advantage of the protease's activity to initiate a cytotoxic cascade, this approach is thought to be less susceptible to known resistance mechanisms.
Laboratori d'Enginyeria de Proteïnes, Departament de Biologia, Universitat de Girona, Girona, Spain.
Mariona
M
Callís
Soraya
S
Serrano
Antoni
A
Benito
Douglas V
DV
Laurents
Maria
M
Vilanova
Marta
M
Bruix
Marc
M
Ribó
D000595:Amino Acid Sequence; D004795:Enzyme Stability; D015658:HIV Infections / Q000201:enzymology / Q000473:pathology; D016333:HIV Protease / Q000737:chemistry* / Q000235:genetics; D015497:HIV-1 / Q000737:chemistry; D006790:Host-Parasite Interactions; D006801:Humans; D007239:Infections / Q000201:enzymology / Q000473:pathology; D009682:Magnetic Resonance Spectroscopy; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012260:Ribonucleases / Q000737:chemistry* / Q000235:genetics
D012260:Ribonucleases; D016333:HIV Protease; C514803:p16 protease, Human immunodeficiency virus 1; C058703:ranpirnase