Kinetics and mechanism of autoprocessing of human immunodeficiency virus type 1 protease from an analog of the Gag-Pol polyprotein.
AUTOR(ES)
Louis, J M
RESUMO
Upon renaturation, the polyprotein MBP-delta TF-Protease-delta Pol, consisting of HIV-1 protease and short native sequences from the trans-frame protein (delta TF) and the polymerase (delta Pol) fused to the maltose-binding protein (MBP) of Escherichia coli, undergoes autoprocessing to produce the mature protease in two steps. The initial step corresponds to cleavage of the N-terminal sequence to release the protein intermediate Protease-delta Pol, which has enzymatic activity comparable to that of the mature enzyme. Subsequently, the mature enzyme is formed by a slower cleavage at the C terminus. The rate of increase in enzymatic activity is identical to that of the appearance of MBP-delta TF and the disappearance of the MBP-delta TF-Protease-delta Pol. Initial rates are linearly dependent on the protein concentration, indicating that the N-terminal cleavage is first-order in protein concentration. The reaction is competitively inhibited by pepstatin A and has a pH rate profile similar to that of the mature enzyme. These results and molecular modeling studies are discussed in terms of a mechanism in which a dimeric full-length fusion protein must form prior to rate-limiting intramolecular cleavage of the N-terminal sequence that leads to an increase in enzymatic activity.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=44526Documentos Relacionados
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