Carboxypeptidase A: Difference between revisions

In the same way that the S1' subsite is involved in anchoring the polypeptide substrate in place, the <scene name='69/694222/3cpas1subsitespacefill/3'>S1 subsite</scene> (spacefill view, subsite in magenta) contains several residues that help hold the substrate in the active site, but the S1 subsite also contains the residues that are involved in the catalytic chemical mechanism.  In general, the residues of the <scene name='69/694222/3cpas1subsitemeshfill/2'>S1 subsite</scene> (pseudo-mesh view, subsite in magenta) have polar or charged side chains that either allow for hydrogen bonding to stabilize negatively charged intermediates of the hydrolysis reaction or position particular atoms appropriately to allow for chemistry to occur.  Three residues (<scene name='69/694222/3cpas1subsiteresidues1/3'>Asn144, Arg145, and Tyr248</scene>) aid in the recognition of the C-terminal residue of a polypeptide substrate.<ref name="CPA1" />  The Asn144 and Tyr248 residues each engage in [http://en.wikipedia.org/wiki/Hydrogen_bond hydrogen bonding] and [http://en.wikipedia.org/wiki/Intermolecular_force ion-dipole interactions] with the carboxyl group at the C-terminus, while Arg145 provides additional stability by participating in [http://www.masterorganicchemistry.com/2010/10/01/how-intermolecular-forces-affect-boiling-points/ ion-ion interactions] with the carboxyl group (see Figure 3 in the section titled "Mechanism of Action").  <scene name='69/694222/3cpas1subsiteresidues2/2'>Arg71</scene> helps stabilize the substrate in the active site by engaging in ion-dipole interactions with the carbonyl oxygen of the penultimate substrate residue (Figure 3).  Three residues (<scene name='69/694222/3cpas1subsitezn/3'>His196, Glu72, and His69</scene>) are liganded to a catalytic Zn²⁺ ion that is complexed to a water molecule positioned one bond distance away from the C-terminal peptide bond carbonyl carbon (Figure 3).<ref name="CPA2" />  <scene name='69/694222/3cpas1subsiteglu270/3'>Glu270</scene> deprotonates this water molecule and acts as a base catalyst in the hydrolysis mechanism (Figure 3).  <scene name='69/694222/3cpas1subsitearg127/3'>Arg127</scene>, along with the positively charged Zn²⁺ ion, help stabilize the negatively charged intermediate generated in the [http://www.masterorganicchemistry.com/tips/addition-elimination/ addition-elimination] step of the hydrolysis reaction (Figure 3).<ref name="CPA2" />

As previously stated, <scene name='69/694222/1cpx_default/3'>CPA</scene> from ''B. taurus'' has the ability to bind two Zn²⁺ ions in its active site.  The binding of only one Zn²⁺ ion is [http://en.wikipedia.org/wiki/Catalysis catalytic], while the binding of a second is [http://en.wikipedia.org/wiki/Reaction_inhibitor inhibitory].  These Zn²⁺ ions are connected to each other via a hydroxy-bridge (Figure 4) with a distance of 3.48 [http://en.wikipedia.org/wiki/%C3%85ngstr%C3%B6m Å] and in tetrahedral arrangement.<ref name="CPA1" />   In the CPA structure containing only the catalytic Zn²⁺ ion (3CPA), a water molecule complexed to the zinc is able to be deprotonated by <scene name='69/694222/3cpas1subsiteglu270/3'>Glu270</scene> to allow for normal initiation of hydrolysis.  Again, this water molecule was not crystallized in the structure of 3CPA, but it is shown in Figure 3.  However, when <scene name='69/694222/Glu270wiz/8'>the inhibitory zinc ion</scene> is also present ([http://www.rcsb.org/pdb/explore/explore.do?structureId=1cpx 1CPX]), it occupies the physical space that would normally be occupied by the water molecule.  Thus, the inhibitory Zn²⁺ ion interacts with the carboxylate group of Glu270.  The Glu270 (shown in yellow) now simply stabilizes the second Zn²⁺ and is unable to perform its usual base catalyst role while the catalytic Zn²⁺ ion (shown in cyan) is still being stabilized in place by His69, Glu72, and His196 (shown in orange).