MF7000279 Beta-carbonic anhydrase CafC (L25G and L78G mutant) 6jqd X-ray 1.69 Homodimer Aspergillus fumigatus 31376470 Kim S, Kim NJ, Hong S, Kim S, Sung J, Jin MS J. Struct. Biol. 2019 1 208 61-68 The β-carbonic anhydrases (β-CAs) are widely distributed zinc-metalloenzymes that play essential roles in growth, survival, development and virulence in fungi. The majority of filamentous ascomycetes possess multiple β-CA isoforms among which major and minor forms have been characterized. We examined the catalytic behavior of the two minor β-CAs, CafC and CafD, of Aspergillus fumigatus, and found that both enzymes exhibited low CO₂ hydration activities. To understand the structural basis of their low activities, we performed X-ray crystallographic and site-directed mutagenesis studies. Both enzymes exist as homodimers. Like other Type-I β-CAs, the CafC active site has an "open" conformation in which the zinc ion is tetrahedrally coordinated by three residues (C36, H88 and C91) and a water molecule. However, L25 and L78 on the rim of the catalytic entry site protrude into the active site cleft, partially occluding access to it. Single (L25G or L78G) and double mutants provided evidence that widening the entrance to the active site greatly accelerates catalytic activity. By contrast, CafD has a typical Type-II "closed" conformation in which the zinc-bound water molecule is replaced by aspartic acid (D36). The most likely explanation for this result is that an arginine that is largely conserved within the β-CA family is replaced by glycine (G38), so that D36 cannot undergo a conformational change by forming a D-R pair that creates the space for a zinc-bound water molecule and switches the enzyme to the active form. The CafD structure also reveals the presence of a "non-catalytic" zinc ion in the dimer interface, which may contribute to stabilizing the dimeric assembly. GO:0004089 carbonate dehydratase activity GO:0008270 zinc ion binding 2 1 Homooligomeric enzymes Homodimeric enzymes All chains according to the most probable oligomerization state stored in PDBe were considered. A Carbonic anhydrase Aspergillus fumigatus Q4WPJ0 1 164 99% MTNVADIEAANAQYAAAFTKGHLPLPPKRKLAVVTCMDARIDVFSVLGLTEGDAHVIRNAGGRASEALRSLIISQRLLGTEEVVVIHHTDCGMLTFSDEDIRAKIREELGEDASDIKFLPFRDLEASVREDVRFLRGSRLVQGNVTGYVYEVERGRLVRLDVSDI 165 secondary structure helix 2 18 secondary structure helix 42 48 secondary structure helix 63 79 secondary structure helix 91 96 secondary structure helix 97 110 secondary structure helix 123 138 secondary structure strand 31 36 secondary structure strand 54 59 secondary structure strand 82 88 secondary structure strand 144 150 secondary structure strand 157 160 pfam PF00484 Carbonic anhydrase 32 99 B Carbonic anhydrase Aspergillus fumigatus Q4WPJ0 1 164 99% MTNVADIEAANAQYAAAFTKGHLPLPPKRKLAVVTCMDARIDVFSVLGLTEGDAHVIRNAGGRASEALRSLIISQRLLGTEEVVVIHHTDCGMLTFSDEDIRAKIREELGEDASDIKFLPFRDLEASVREDVRFLRGSRLVQGNVTGYVYEVERGRLVRLDVSDI 165 secondary structure helix 2 17 secondary structure helix 42 48 secondary structure helix 63 79 secondary structure helix 91 95 secondary structure helix 97 110 secondary structure helix 123 138 secondary structure strand 31 36 secondary structure strand 54 59 secondary structure strand 82 88 secondary structure strand 144 150 secondary structure strand 157 160 pfam PF00484 Carbonic anhydrase 32 99 Indirect evidence The full structure participates in mutual synergistic folding. Carbonic anhydrase The native carbonic anhydrase is dimeric in solution, in agreement with being a tightly associated dimer with a large, hydrophobic buried surface area. The extended β-sheet core consisting of ten β–strands is equally contributed by the two monomers, and the N-terminal helix of each monomer extends around the other monomer. Based on the highly intertwined structure, the monomeric form is most probably not a stable, independently folding unit. The two active sites are also located in clefts at the dimeric interface, further suggesting that dimer is the functional form (PMID:32515610). A N/A B N/A MF7000278 MF7000279 MF7000856 MF7000857 MF7000858 MF7000859