Figure 6
V86M CA interactions with CypA studied by NMR. (A) Expansions of 2D 1H-15N ZZ-exchange spectra illustrating CypA catalysed cis-trans proline isomerisation of P90 in HIV1caN. The four panels represent backbone amide 1H,15N correlations for the preceding G89 residue in HIV1caN and in HIV1caN V86M in the presence and absence of CypA. In the absence of CypA, the isomerisation reaction is slow and only distinct G89 cis- and trans- correlation peaks are observed. The presence of CypA leads to an increase in the isomerisation rate and the accumulation of cis and trans exchange peaks. The dotted box denotes the location of the exchange peaks and indicates their symmetrical distribution with regard to G89 cis and G89 trans . Note that in the case of HIV1caN V86M a broad G89cis peak and an additional cis-exchange peak indicates a second conformer. All spectra were acquired at the same mixing time (69 ms) and processed with the same contour levels. (B) Evaluation of exchange rates for CypA catalyzed isomerisation of G89-P90 in HIV1caN and HIV1caN V86M. Normalised peak intensities of trans auto peaks and the corresponding exchange peaks for HIV1caN/CypA and HIV1caN V86M/CypA were plotted as a function of mixing time (s). Exchange rates (kex) in the order of 5 s-1 for HIV1caN/CypA and 20 s-1 for HIV1caN V86M/CypA were extracted by fitting auto peaks and exchange peaks as described by Bosco et al. (2002).