## 5 CO\({}_{2}\)/O\({}_{2}\) Molecular Recognition Strategy  
In the same study [15], [PERSON] was able to propose a mechanistic explanation for the Rubisco ability to discriminate between CO\({}_{2}\) and O\({}_{2}\) at \"RuBP-binding site\". As intracellular (CO\({}_{2}\)) is estimated approximately 7-8 uM in C3 plants and 80 uM in C4 plants, while [O\({}_{2}\)] is about 260 uM, [30] a key point in Rubisco catalysis is the CO\({}_{2}\)/O\({}_{2}\) competition for the RuBP-binding active site. To compensate for the low CO\({}_{2}\) intracellular level, Rubisco shows K\({}_{\rm C}\) values ranging from 8 to 19 uM and K\({}_{\rm O}\) values ranging from 230 to 620 uM while S\({}_{\rm C/O}\) values range between 70 and 100 in plants (considering both C3 e C4 plants) and can reach values as high as 222 in _Galderia subhuraria_ (a red algae). Considering that Rubisco does not have specific binding sites for CO\({}_{2}\) or O\({}_{2}\) and the two gaseous molecules are not so different as far as dimensions and electrostatic potential surfaces are concerned (Figure 6a), the Rubisco higher specificity for CO\({}_{2}\) is not easily explained. Stec [15] proposes that the two gaseous molecules \"bind\" to the same region of the active site, at the mouth of the TIM barrel, covered as a lid by the N-terminal domain of the adjacent L subunit (see Figure 3), where a high electrostatic field gradient is set up by the closeness of negative and positive surfaces potential. Figure 6b shows a negative surface potential (represented in red) at the N-terminal domain and a positively charged cavity (represented in blue) at the C-terminal domain. As the quadrupole moment of CO\({}_{2}\) is approximately 15-fold higher than that of O\({}_{2}\) (for CO\({}_{2}\) Qzz = \(-\)3.239 e a\({}_{0}\)\({}^{2}\), for O\({}_{2}\) Qzz = \(-\)0.232 e a\({}_{0}\)\({}^{2}\) as calculated by Stec), the quadrupole mo  
Figure 5: Mechanism of the Rubisco activation reaction as proposed by [PERSON]. (**a**) Shows the arrangement of the His335 side chain contributing to the “activation binding site”; (**b**) shows that the His-335 side chain swings out to an alternative conformational state allowing a Mg\({}^{2+}\) ion binding; (**c**) after proton abstraction, Lys201 swings in a position close to the CO\({}_{2}\)-carbon affording the carbamylated-Lys201. The gray arrow shows the second OH\({}^{-}\) group abstracting H\({}^{+}\) from the NH\({}_{2}\)-carbamyl-moiety; (**d**) formation of a Mg\({}^{2+}\)-stabilized acid-carbamate complex reaching a new location and prone to start the catalytic reactions (Figure from ref. [15], used by permission of PNAS (copyright © 2012)). ligands interacting with the very strong electric field built across the active site allows discrimination between the two species. The hypothesis advanced by [PERSON] appears correct and provided a rationale for the minimal requirement of a L\({}_{2}\) dimer as a basic functional unit found across the Rubisco protein family. In fact, contact between the surfaces of the two subunits (facing each other) is needed to build up the gradient of the electric potential. [PERSON] et al. (2018) have also faced the problem concerning the favourable competition of CO\({}_{2}\) vs. O\({}_{2}\) by investigating, via molecular dynamics simulations, the interaction of the two gaseous molecules with the protein. A modelled L\({}_{8}\)S\({}_{8}\) hexa-decamer was sol  
Figure 6: (**a**) Computed electrostatic potential molecular surfaces of CO\({}_{2}\) (left) and O\({}_{2}\) (right). The color scheme follows commonly accepted conventions: blue, positive; red, negative. The value of the Qzz component of the quadrupole moment, as calculated by Stec, is \(-3.239\) e a\({}_{0}\)\({}^{2}\) for CO\({}_{2}\) and \(-0.232\) e a\({}_{0}\)\({}^{2}\) for O\({}_{2}\); (**b**) a ribbon representation of the catalytic domain with bound gaseous ligands and surfaces colored by the electrostatic potential. O\({}_{2}\) (in red) and CO\({}_{2}\) (in purple) lie in a positively charged cavity (blue) of the TIM barrel.