5FQK: W229d And F290w Mutant Of The Last Common Ancestor Of Gram-negative Bacteria (gnca4) Beta-lactamase Class A Bound To 5(6)- Nitrobenzotriazole (ts-analog)

Protein engineering studies often suggest the emergence of completely new enzyme functionalities to be highly improbable. However, enzymes likely catalysed many different reactions already in the last universal common ancestor. Mechanisms for the emergence of completely new active sites must therefore either plausibly exist or at least have existed at the primordial protein stage. Here, we use resurrected Precambrian proteins as scaffolds for protein engineering and demonstrate that a new active site can be generated through a single hydrophobic-to-ionizable amino acid replacement that generates a partially buried group with perturbed physico-chemical properties. We provide experimental and computational evidence that conformational flexibility can assist the emergence and subsequent evolution of new active sites by improving substrate and transition-state binding, through the sampling of many potentially productive conformations. Our results suggest a mechanism for the emergence of primordial enzymes and highlight the potential of ancestral reconstruction as a tool for protein engineering.
PDB ID: 5FQKDownload
MMDB ID: 145743
PDB Deposition Date: 2015/12/11
Updated in MMDB: 2016/12
Experimental Method:
x-ray diffraction
Resolution: 1.77  Å
Source Organism:
Similar Structures:
Biological Unit for 5FQK: monomeric; determined by software (PISA)
Molecular Components in 5FQK
Label Count Molecule
Protein (1 molecule)
Gnca4 Lactamase W229d and F290w
Molecule annotation
Chemical (1 molecule)
* Click molecule labels to explore molecular sequence information.

Citing MMDB