CC Chemokine Receptor Type
2 (PDB ID: 5T1A) from Homo
sapiens
Created by: John Kanu
CC Chemokine Receptor Type 2 (PDB ID: 5T1A), as
found in Homo sapiens, is a transmembrane signaling protein which
acts as a chemokine receptor. Chemokine receptors are present in all vertebrate
and are part of the superfamily of class A seven-transmembrane guanine
nucleotide-binding protein, or G protein-coupled receptors (GPCRs) (1). There
are twenty-three known human chemokine receptors, found in human tissue and
leukocytes. They are divided into two groups, GCPR receptors and atypical
receptors. CCR2 is expressed in its physiological form in monocytes, immature
dendritic cells, T-cells, and migrate to the Monocyte Chemoattractant Protein,
MCP-1, found in the endogenous CC Chemokine Ligand known as, CCL2 (PDB ID:
2LIE). CCL2 is synthesized within the splenic arteriolar lymphatic sheet
and medullary regions of the lymph node and other tumors (2). CCR2 bound to
CCL2 and associated ligands are connected to numerous inflammatory and
neurodegenerative diseases including atherosclerosis, multiple sclerosis,
asthma, neuropathic pain, diabetic nephropathy, as well as cancer.
Knowledge of the associations has allowed for studies and trials to develop
preventative therapies that target the CCR2-chemokine receptor. Finding the
proper drug therapies is done by studying the structure in ternary complexwith orthosteric (BMS-681) (PDB ID: 73R) and allosteric antagonists (CCR2-RA-[R]) (PDB ID: VT5)(1).
Through
lipidic cubic phase of the ternary complex (fig. 1) of CCR2, BMS-681, and CCR2-RA-[R].
CCR2 has the capability to form. Other crystals of importance that form are
crystals of CXC chemokine receptor type 4 (PDB ID: 30E6, 3O30, 30DU). The secondary structure of CCR2
is comprised of 62% alpha-helices (23 helices, 316 residues), 5% beta-sheets (6
strands; 27 residues), and 33% random coils (165 residues), and contains a single A chain subunit composed of 508 residues (3). It has a molecular weight
of 57805.54 Da and an isoelectric point (pI) of 9.30 (4). The central fold of
CCR2 contains seven transmembrane alpha-helices connected by three
extracellular and intracellular loops (5). The first two extracellular domains
and loops do not serve any functional significance. The N-terminal domain is
for ligand binding, and the C-terminal domain is towards the cytoplasm. CCR2
shares structural properties that are similar among other chemokine receptors.
The extracellular portion is net-negative under normal physiological
conditions, while the intracellular portion near the C-terminus has serine and
threonine residues that phosphorylate due to Dynamin, a G protein receptor
kinase, GRK. Phosphorylation results in the uncoupling of the G protein
subunits from the receptor. The phosphorylation and GRK recruit
adaptor molecules that link the receptor to a lattice of clathrin that
facilitates receptor internalization (6). Ultimately,
chemokine receptors are identified by a short N-terminus, a short basic third
intracellular domain and a cysteine in each of the three extracellular domains
(1).
Additional structural features distinguish CCR2 from other chemokine receptors. CCR2 binds to other important ligands: (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate, zinc ion, and sulfate (1). CCR2 has two important ionic bonds. First, zinc, a metal ion, binds the side chains together. γ-lactam secondary exocyclic amine forms a hydrogen bond with the hydroxyl of His-144,Thr-292, Glu-238, and Glu-1005. There are four prosthetic ligands, VT5 and 73R, and OLC and the Sulfate Ion. (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate contains two active alcohol groups and allows for crystallization. The sulfate ion (SO42-) helps define the substrate-binding site. CCR2 uses the sulfate to bind to the MCP-1 (2).
The function of the subunit is due to the addition of an orthosteric ligand BMS-681 and allosteric CCR2-RA-[R] antagonist ligand which leads to a site overlap which allows for a highly favorable drug pocket. This drug pocket that
is created is highly due to the allosteric CCR2-RA-[R] antagonist. CCR2-RA-[R]
is surrounded by a cage using the intracellular ends of helices I-III and
VI-VIII. The mixed polarity of CCR2 creates a hydrophobic inner area is surrounded by residuesVal-63, Leu-67,
Leu-81, Leu-134, Ala-241, Val-244, Tyr-305, and Phe-312. The outer polar
portion of the cage is comprised of Thr-77, Arg-318, Glu-309, Lys-311, and
Tyr-315. There is a backbone of residues Arg-237 and Lys-240 (1,2). This binding
pock is tightly enclosed and is balanced between hydrophobic and polar, which
allows for the site to be highly druggable. The binding bock is created in part
of two disulfide bonds, between Cys-32- Cys-277 and Cys-113-Cys-190.
CCR2-RA-[R] in its natural form sterically hinders CCR2; therefore, when BMS-681
binding interferes with the chemokine binding site directly and with the GPCR
indirectly, forcing an incompatible conformation. Natural receptor-chemokine
interactions prove to be challenging for therapy, as the CCR2 has selectivity
for multiple ligands which block the ability for a drug pocket (6).
Pharmaceutical antagonist and orthosteric ligands inhibit the binding of CCR2
to CCL2, allowing for drug pocket availability (2).
CCR2
shows structural similarities to the Mu-type opioid receptor, lysozyme chimaera(PDB ID: 4DKL) found within Mus musculus. Mu-type opioid receptor
is comprised of 70% alpha-helices (20 helices, 326 residues), 4% beta-sheets (8
strands; 28 residues), and 26% random coils (110 residues), and contains a single A chain subunit composed of 464 residues. It has a molecular weight
of 55761.68 Da and an isoelectric point (pI) of 9.37 (4,7). Comparisons
between the Mu-type opioid receptor and CCR2 resulted in a BLAST E-score of 2.0
x 10-177 (8). An E-score of less than 0.05 indicates that the two proteins have
a similar primary structure. The DALI server generated a Z-score of 30.8 (9). A
Z-score more significant than 2 indicates that the two proteins are similar
among their tertiary structure. The Mu-type opioid receptor binds to six
associated ligands. One that is similar to that of CCR2, which is a sulfate ion. The other associated ligands involve: a chloride ion which acts as an
ionic bond binding the side chains of the receptor: Ser-261, Glu-270, and Lys-271; [(Z)-octadec-9-enyl]
(2R)-2,3-bis(oxidanyl)propanoate, whose role is to act as an intermediate; an
antagonist ligand methyl
4-{[(5beta,6alpha)-17-(cyclopropylmethyl)-3,14-dihydroxy-4,5-epoxymorphinan-6-yl]amino}-4-oxobutanoate;
a water-soluble, Pentaethylene glycol and cholesterol who modulates the
functions of the protein. Mu-type opioid receptor (55761.68 Da) differs from CC
Chemokine Receptor 2 (59577.95 Da) by mass, polarity, as well as the multitude (11:4) of
extra sulfate ions present. There are also more ligands present in Mu-type
opioid receptor than in CC Chemokine Receptor 2 (7).
CCR2
can conform to two similar structures that are not just crystallized versions
of itself. First, there is the crystal structure of the CCR4 chemokine receptor
in complex with a small molecule antagonist IT1t in I222 space-group (PDB ID:
3OE6). Then there is the crystal structure of CXCR4 and viral chemokine
antagonist MIP-II complex (PSI Community Target) (PDB ID: 4RWS) (10). Duchenne
muscular dystrophy is the progressive weakness of muscle that is often fatal
due to the loss of control of the diaphragm. A dual chemokine receptor, CCR2/CCR5 (PDB ID: 4MBS), treated with 20mg/kg/day of cenicriviroc, an
antagonist, can prevent the macrophage accumulation and the progression of the
disease (11). CCR2 is a complex receptor protein that can lead to detrimental
effects on the human body if not altered with BMS-681 and CCR2-RA-[R].
The effect of the alteration prevents the CCR2 from binding to CCL2, which has
the possibility of leading to neuromuscular disease and cancers within the
human body (1).