The solution structure of VAT-N reveals a ‘missing link’ in the evolution of complex enzymes from a simple βαββ element

Results: Using nuclear magnetic resonance (NMR) spectroscopy, we found that VAT-N is composed of two equally sized subdomains. The amino-terminal subdomain VAT-Nn (comprising residues Met1–Thr92) forms a double-psi β-barrel whose pseudo-twofold symmetry is mirrored by an internal sequence repeat of 42 residues. The carboxy-terminal subdomain VAT-Nc (comprising residues Glu93–Gly185) forms a novel six-stranded β-clam fold. Together, VAT-Nn and VAT-Nc form a kidney-shaped structure, in close agreement with results from electron microscopy. Sequence and structure analyses showed that VAT-Nn is related to numerous proteins including prokaryotic transcription factors, metabolic enzymes, the protease cofactors UFD1 and PrlF, and aspartic proteinases. These proteins map out an evolutionary path from simple homodimeric transcription factors containing a single copy of the VAT-Nn repeat to complex enzymes containing four copies.