Abstract:
:The reverse transcriptase (RT) from the human immunodeficiency virus (HIV) is initially expressed as a 66-kDa protein and is subsequently proteolytically processed in vivo to form a 66-kDa/51-kDa heterodimer. Comparison of circular dichroism spectra of the 66-kDa, 51-kDa, and heterodimeric forms of RT indicates that the conversion is accompanied by dramatic changes in subunit conformation. The mean residue ellipticity per subunit at 220 nm decreases from -10.7 x 10(3) deg cm2 dmol-1 for the 66-kDa protein to -6 x 10(3) deg cm2 dmol-1 for the heterodimer. The same loss of ellipticity is observed whether the heterodimer is produced by proteolysis or by mixing a separately-expressed cloned 51-kDa subunit with the 66-kDa protein. Comparison with the spectrum of the cloned 51-kDa protein suggests that much of the conformational change arises from formation of the 51-kDa subunit but substantial changes occur in the remaining 66-kDa subunit as well. A kinetic analysis was performed to correlate these conformational changes with changes in enzyme function. Application of an integrated Michaelis-Menten equation to the catalysis of poly(dT) formation using a d(pT)20-poly(rA) primer-template shows that the kcat for the heterodimer is approximately half that of the 66 kDa enzyme, decreasing from 2.9 to 1.2 nucleotides/s upon formation of the heterodimer. However, km values for the primer-template decrease from 0.54 to 0.12 microM upon heterodimer formation. Thus, kcat/Km is 2-fold larger for the heterodimer, giving it a distinct catalytic advantage at undersaturating concentrations of enzyme and primer-template.(ABSTRACT TRUNCATED AT 250 WORDS)
journal_name
Biochemistryjournal_title
Biochemistryauthors
Anderson SF,Coleman JEdoi
10.1021/bi00150a015subject
Has Abstractpub_date
1992-09-08 00:00:00pages
8221-8issue
35eissn
0006-2960issn
1520-4995journal_volume
31pub_type
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