A metalloenzyme known as thermolysin is inhibited by a compound known as Structure IX. There are two analogues synthesized for Structure IX, and they both substitute the functional group interacting with the carbonyl oxygen atom of the enzyme active site. One analogue has similar binding affinity compared to the original Structure IX, while the other analogue has reduced binding affinity.
Structure IX has a secondary amine functional group (NH) that interacts with the carbonyl oxygen atom of the thermolysin active site. This interaction occurs via hydrogen bonding.
The electronegative nitrogen atom of the secondary amine gives the hydrogen atom of the amine group a partial positive charge, allowing it to attract the lone pair of electrons belonging to the oxygen atom in the thermolysin active site. This type of bond is known as a hydrogen bond.
However, one analogue substitutes the secondary amine (NH) for a single oxygen atom (O). This eliminates the hydrogen bonding between the enzyme and the compound, significantly weakening its interaction. Hence, the analogue has reduced affinity by a factor of 1000 due to the removal of the hydrogen bonding groups.
Another analogue, however, replaces the secondary amine (NH) with a methylene (CH2) group. This methylene group is capable of hydrogen-bonding with the carbonyl oxygen atom in the enzyme active site, despite the slightly lower electro-negativity of the carbon atom.