Advances in the field of recombinant protein engineering and expression, protein purification, NMR, X-ray crystallography and computational chemistry have improved the skills of drug designers to use data that have been gathered on the three-dimensional structures of protein ligand complexes.
Most bacterial species have various types of PBP which differ in various ways such as enzymatic function, molecular weight and the affinity for β-lactam antibiotics. There are two types of enzymes that are particularly interesting with regard to the binding site of β-lactams: PBP and β-lactamases. Target alterations in the binding site of PBP have led to high-level resistance of β-lactams among bacteria like staphylococci, enterococci and pneumococci.
For example, the binding site of PBP2 in Neisseria gonorrhoeae has been structurally determined and has three sequence motifs that can be seen in nearly all β-lactam interacting enzymes:
* SXXK motif is located at the N-terminal end of α2 helix and includes two residues that are important for the enzyme function.
** Ser-310 : Includes a serine nucleophile that is acylated by both peptide substrate and β-lactam antibiotics.
** Lys-313 : Plays an important role in providing the dense hydrogen bound network at the active site and is in distance of Ser 310, ASN-364 and the carbonyl backbone of Ser-362.
* SXN motif that includes Ser-362, Ser-363 and Asn-364
* KTG motif that includes Lys-497, Thr-498 and Gly-499
Research also implies that adjacent regions to the active site which differ between different PBP have significant influence on the rate of β-lactam acylation rate.