Carboxypeptidase A: Difference between revisions

=====S1 Subsite=====

In the same way that the S1' subsite is involved in anchoring the polypeptide substrate in place, the <scene name='69/694222/3cpas1subsitespacefill/2'>S1 subsite</scene> (spacefill view, subsite in cyan) 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/1'>S1 subsite</scene> (pseudo-mesh view, subsite in cyan) 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='69/694222/3cpas1subsiteresidues1/1'>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 2 in the section titled "Mechanism of Action").  <scene name='69/694222/3cpas1subsiteresidues2/1'>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 2).  Three residues (<scene name='69/694222/3cpas1subsitezn/1'>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 2).<ref name="CPA2" />  <scene name='69/694222/3cpas1subsiteglu270/1'>Glu270</scene> deprotonates this water molecule and acts as a base catalyst in the hydrolysis mechanism (Figure 2).  <scene name='69/694222/3cpas1subsitearg127/1'>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 2).<ref name="CPA2" />

=====Putting It All Together=====

For a 3D manipulative applet displaying the essential amino acid residues and their interactions with a bound ligand, <scene name='69/694222/6cpawithligandmesh/1'>click here</scene>.  The hydrophobic binding pocket (shown in yellow) can be seen interacting with a phenylalanine side chain of the C-terminal amino acid.  The Zn²⁺ ion is held in place by the three side chains mentioned previously (shown in green).  The Tyr248 side chain is explicitly shown in cyan and is "capping" the pocket since a substrate is bound.  The base catalyst, Glu270, is explicitly shown in orange.  The other side chains in the darker blue color are part of the stabilizing side chains of the S1 subsite.  Manipulate the scene manually, and zoom in and out to see the interactions between CPA's side chains and the bound substrate.  This crystal structure with the bound ligand, which is actually a [http://en.wikipedia.org/wiki/Phosphonate phosphonate], has been deposited in the Protein Data Bank (PDB) with code [http://www.rcsb.org/pdb/explore/explore.do?structureId=6CPA 6CPA].

== Mechanism of Action ==