home - index - site map - plugin if you don't see a molecular model:
http://www.papanatur.de/fett/fabf/fabfm.htm

FabF, a beta-ketoacyl-ACP synthase II from Escherichia coli : inhibition by platensimycin

Structure - The protein chain is folded into distinct subunits. The aminoterminal part (residues 1-251) has a central beta sheet of mixed topology. The sheet is flanked by helices  . The carboxyterminal subunit has a quite similar structure making up for a alpha-beta-alpha-beta-alpha sandwich for the whole chain  .

Catalytic center - The enzymatically active amino acid is Cys¹⁶³ which is at the bottom of a pocket lined by highly conserved mostly hydrophobic residues  . You may notice the odd shape of the protein in this view and the edgy position of the catalytic center. In reality it looks much better, as there is the second chain of the dimeric enzyme  . Some amino acids of the other protein are part of the hydrophobic channel too (the dimer being symmetric, you find a second catalytic center on the other side as well). In the catalytic center two histidines are essential for the fatty acid condensation step. His³⁴⁰ functions in catalysis by abstracting a proton from Cys¹⁶³. Another obvious feature is the inaccessibility of the catalytic center to a bulky substrate. Phe⁴⁰⁰ seems to block the entry to the bottom of the pocket. If the enzyme is supposed to work, the sidechain of Phe⁴⁰⁰ has to move away during the reaction. A look at a reaction intermediate shows that it does. To visualize this, an experimental trick had to be employed: the reaction intermediate of the wild type enzyme is too unstable to be cought by X-ray analysis. An exchange of Lys³³⁵ to alanine affects the orientations of the essential histidines to an extent that a reaction intermediate becomes visible: a Cys¹⁶³-lauroyl thioester. The carbonyl oxygen of the ester is oriented into the oxyanion hole established by the backbone amide nitrogen of Phe⁴⁰⁰  . The second substrate, malonate which after decarboxylation attacks the thioester, finds place in the upper part of the binding pocket (malonate is not visible in this experiment).

Binding of platensimycin - By labelling experiments the antibiotic platensimycin was found to bind to the beta-ketoacyl-ACP synthase. However, binding depended on an acylated intermediate form of the enzyme. To obtain a well diffracting crystal for X-ray investigations, the enzyme had to be modified to mimick the intermediate. Cys¹⁶³ of the wild type was replaced by glutamine  . The amide oxygen of this substitute resembles the acyl carbonyl oxygen with respect to the oxyanion hole  . This makes the perfect binding stage for platensimycin which neatly fits into the binding pocket  . One oxygen atom of the platensimycin carboxyl group is within hydrogen bridge distance to the catalyc histidines of the enzyme  , the aromatic ring stacks edge on to Phe⁴⁰⁰  . The amide nitrogen bridges to Thr²⁷⁰  , the amide oxygen to Thr³⁰⁷ as does the hydroxyl group in the ring system  . The ether oxygen bridges to Thr²⁷⁰ again  . A water molecule mediates further contacts  . So this sums up to a total blockage of the binding pocket for the second substrate of the enzyme, malonate  .