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[37]. For RuvC, four residues have been shown to be essential for catalysis [40]. The third of these,
Asp138, is at the amino end of the last helix of the structure. In the three-dimensionally aligned
structures, this helix is parallel with the helix D in HIV-1 integrase, although it is running in the opposite
direction. The position of helix D in RuvC is also significantly different, mainly due to a 13 Å shift
along its axis toward the active site, placing Asp138 close to the other two carboxylates. The fourth
essential residue, Asp141, is on the first turn of the same helix, very close in the aligned structures to the
essential Glu157 of ASV integrase, their α carbons separated by only 1.6 Å. For the MuA transposase,
its third essential carboxylate, Glu392, is in a loop region, just one residue upstream from the amino end
of a helix, the topological equivalent of helix D. Unexpectedly, this residue turns away from the region
defined by the two other carboxylates to a position where it clearly cannot contribute to the formation of
a metal-binding site. It is likely, therefore, that the conformation of the polypeptide chain around Glu392
in the transposase core observed in the crystal structure belongs to an inactive form. In this case, a
conformational change upon transposase tetramer assembly or perhaps upon substrate binding is
required for activity.
Significance of the Disordered Region
From the point of view of HIV-1 integrase, it is interesting to note that the apparently flexible part of the
MuA transposase structure is topologically equivalent with the disordered and uninterpretable part of the
integrase. Similarly, in the crystal structure of the isolated RNase H domain of HIV-1 reverse
transcriptase, a five-residue loop in a topologically equivalent location is disordered and therefore
uninterpretable. In the ASV integrase core, the corresponding loop is visible but with rather high
mobility. It seems that some kind of disorder or flexibility in this region is a common feature of the
superfamily. Crystal structures of enzyme-substrate or enzyme-inhibitor complexes will tell us the
functional significance of this flexibility as well as the exact configuration of the active site.
C. The Dimer Interface
HIV-1 integrase is active as a multimer, and the catalytic core domain alone forms dimers in solution,
even at low protein concentration (see Section II.C). In the crystal structure, a roughly spherical dimer of
about 45 Å diameter was observed, formed by a crystallographic two-fold axis. The dimer has a large
solvent-excluded surface of 1300 Å 2 per monomer. This area is close to what is expected for dimers in
this molecular weight range [41]. Therefore, we are convinced that in the crystal structure the authentic
dimer is present. This was subsequently confirmed by the structure of the ASV integrase core. Although
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