**2. The importance and multifaceted functionality of GPCRs**

The importance of G-protein coupled receptors (GPCRs) in the fields of biology, medicine and pharmaceutical studies have been extensively studied, well established and properly documented [7]. Due to its significance in playing a crucial role in various normal and pathological processes, GPCRs have become a major field of advanced research and a promising focus for drug discovery processes. The GPCRs have an extensive medical significance owing to their position and function within the human cell spanning the whole cell's plasma membrane. By this way it bridges the extra- and an intracellular environment which enables the GPCRs to act as signal transducers wherein it acclaims a direct mechanism for the transduction of extracellular messages into intracellular responses. In this way and together with their transmitters and effectors, GPCR systems function to modulate a broad spectrum of cellular phenomena dictated by the needs of the tissues and organs they serve. The gradient of GPCR distribution across vast majority of the body's organs and tissues and its primary role as signal transducers like converting transduce extracellular stimuli into intracellular signals at cellular levels makes it fascinating molecules from the perspective of advanced structural research.

Other fascinating roles of GPCRs include modulation of neuronal firing, regulation of ion transport across the plasma membrane and within intracellular organelles, modulation of homeostasis, control of cell division/proliferation, and modification of cell morphology. GPCRs are also an important target for cardiac drug therapy as decades of research revealed that GPCRs are the epicenters of many of the multiple causative factors of cardiovascular diseases like diabetes, obesity, environmental stressors and genetic factors [8]. Thus understanding the GPCR signaling mechanism in a healthy and an ailing heart may give better insights into treating cardiovascular problems.

There are over 200 cardio GPCRs and understanding their structural and functional properties is a key element in understanding the occurrence of heart diseases [9]. G-proteins consist of α, β, and γ subunits and a lot of global research has been carried out to check the various GPCR signaling pathways in a healthy and an ailing heart. Clinically targeted cardiac GPCRs like adrenergic receptors are responsible for translating chemical messages from the sympathetic nervous system into cardiovascular responses. Other such potentially targeted clinical GPCRs include angiotensin, endothelin, and adenosine receptors. Thus to study deeper about such cardio GPCRs one has to have structural studies carried out prior to analyzing its functionality.

Chemokine receptors belonging to the class A of GPCRs are involved in variety of physiologic functions, mostly related to the homeostasis of the immune system. They are also involved in multiple pathologic processes, including immune and autoimmune diseases, as well as cancer.

Other ailments caused when fundamental pathways governed by GPCRs go awry are asthma and strokes and cerebral hypoperfusion [10]. GPCRs control airway smooth muscle (ASM) contraction and increased airway resistance when coupled to Gq receptors. Airway epithelium and hematopoietic cells that are involved in control of lung inflammation that causes most asthma, have various pathways that are mediated by GPCRs. Arrestins regulate GPCR signaling and once again structural insights into the GPCRs is essential in understanding vital role of arrestins in those GPCR-mediated airway cell functions that are dysregulated in asthma.

## **3. A brief history on the structural study of GPCRs**

GPCRs have been considered as one of the most desirable drug targets for the past few decades and have been investigated extensively. But the three dimensional structures of GPCRs have only recently become available. The first step in the structural study of GPCRs happened in the year 2000 with the initial crystal structure determination of Bovine rhodopsin (PDB: 1F88) through X-ray diffraction method [11]. The GPCR rhodopsin was purified from bovine rod outer segment (ROS) membranes. Multiwavelength anomalous diffraction (MAD) methods were employed to get the phasing information and the diffraction data from the crystallized Bovine rhodopsin were collected to 2.8 Å after mercury soaking. This experimental model of rhodopsin became a structural template for other GPCRs owing to the molecular size of Bovine rhodopsin, 348 amino acids, which was intermediate among the members of the GPCR family and thus can feature most of the essential parts of functional importance in G-protein activation.

An year later in 2001 the solution NMR method was used to solve the structure of Bovine rhodopsin (PDB ID: 1JFP [12]). It then took 7 long years to crystallize the next GPCR ADRB2 (PDB ID: 2RH1, 2R4R/2R4S [13, 14])[15]. It was solved using the LCP method that provides a more native, lipid environment for crystallization to a resolution of 2.4 Å. This delay was due to the need of numerous technological advancements required to crystallize membrane proteins like GPCRs. Developments in protein engineering, computational methods like homology

**59**

**Figure 2.**

[32, 33]).

*Importance of Homology Modeling for Predicting the Structures of GPCRs*

homology modeling in the following years of research.

modeling and heterologous protein expressions have accelerated structural determination of GPCRs [16]. In this structure solved via the LCP technique, the proteins are placed in a membrane-like environment where they can diffuse and interact with each other to form crystal lattice contacts on both complementary hydrophobic and hydrophilic regions. These structures served as the template for the other crystal structures that were solved afterwards. Other receptors like H1R, D3R and 5-HT1B belonging to the Rhodopsin family of GPCRs were solved in the following years and served as templates for all the other GPCR structures that were predicted by

Another important subfamily of class A GPCRs with a number of key physiologic roles are the Chemokine receptors [17]. So far (till 2020) only 5 different chemokine receptor complexes have had their crystal structure solved by researchers and they are CXCR4 [18] (PDB IDs: 3ODU, 3OE0, 3OE6, 3OE8, 3OE9, and 4RWS [18, 19]), CCR5 [20] (PDB IDs: 4MBS, 5UIW, 6AKX, and 6AKY [21–23]), US28 [24] (PDB IDs: 4XT1, 4XT3, 5WB1, and 5WB2 [24, 25]), CCR2 [26] (PDB IDs: 5T1A, 6GPS, and 6GPX [26, 27]) and CCR9 [28] (PDB ID: 5LWE [28]). Structure based drug design was the key in solving crystal structures of Chemokine receptors and its potential is reflected by the large amount of ligands found for various chemokine receptors. SBDD methods prove to

In 2011, Kobilka achieved another break-through when he and his team captured

There are various databases available exclusively for GPCR structures like GPCR-EXP [https://zhanglab.ccmb. med.umich.edu/GPCR-EXP/] (database for experimentally solved GPCR structures) and GPCRdb [34] (web tools and diagrams that aid GPCR research) that profusely help the researchers. According to GPCR-EXP statistics there are 389 structures for 67 GPCRs belonging to different species deposited in the PDB. **Figure 2** gives us details about the total number of new experimental

*The total number of new GPCR experimental structures available each year given by GPCR EXP database.*

an image of the β-adrenergic receptor at the exact moment that it is activated by a hormone and sends a signal into the cell. This image is a molecular masterpiece [29]. This was the first step in the path that earned Brian Kobilka the Nobel Prize in Chemistry in the year 2012 for his groundbreaking discoveries about GPCRs along with Robert Lefkowitz. In the year 2011 and 2013, the first secretin family GPCR structure was solved (PDB ID: 4L6R, 4K5Y [30, 31]) and in the following year the first glutamate family GPCR structure was deposited in PDB (PDB ID: 4OR2, 4OO9

be more effective when a crystal structure is available as homology models.

*DOI: http://dx.doi.org/10.5772/intechopen.94402*

#### *Importance of Homology Modeling for Predicting the Structures of GPCRs DOI: http://dx.doi.org/10.5772/intechopen.94402*

*Homology Molecular Modeling - Perspectives and Applications*

treating cardiovascular problems.

autoimmune diseases, as well as cancer.

functionality.

organelles, modulation of homeostasis, control of cell division/proliferation, and modification of cell morphology. GPCRs are also an important target for cardiac drug therapy as decades of research revealed that GPCRs are the epicenters of many of the multiple causative factors of cardiovascular diseases like diabetes, obesity, environmental stressors and genetic factors [8]. Thus understanding the GPCR signaling mechanism in a healthy and an ailing heart may give better insights into

There are over 200 cardio GPCRs and understanding their structural and functional properties is a key element in understanding the occurrence of heart diseases [9]. G-proteins consist of α, β, and γ subunits and a lot of global research has been carried out to check the various GPCR signaling pathways in a healthy and an ailing heart. Clinically targeted cardiac GPCRs like adrenergic receptors are responsible for translating chemical messages from the sympathetic nervous system into cardiovascular responses. Other such potentially targeted clinical GPCRs include angiotensin, endothelin, and adenosine receptors. Thus to study deeper about such cardio GPCRs one has to have structural studies carried out prior to analyzing its

Chemokine receptors belonging to the class A of GPCRs are involved in variety of physiologic functions, mostly related to the homeostasis of the immune system. They are also involved in multiple pathologic processes, including immune and

Other ailments caused when fundamental pathways governed by GPCRs go awry

GPCRs have been considered as one of the most desirable drug targets for the past few decades and have been investigated extensively. But the three dimensional structures of GPCRs have only recently become available. The first step in the structural study of GPCRs happened in the year 2000 with the initial crystal structure determination of Bovine rhodopsin (PDB: 1F88) through X-ray diffraction method [11]. The GPCR rhodopsin was purified from bovine rod outer segment (ROS) membranes. Multiwavelength anomalous diffraction (MAD) methods were employed to get the phasing information and the diffraction data from the crystallized Bovine rhodopsin were collected to 2.8 Å after mercury soaking. This experimental model of rhodopsin became a structural template for other GPCRs owing to the molecular size of Bovine rhodopsin, 348 amino acids, which was intermediate among the members of the GPCR family and thus can feature most of the essential

An year later in 2001 the solution NMR method was used to solve the structure of Bovine rhodopsin (PDB ID: 1JFP [12]). It then took 7 long years to crystallize the next GPCR ADRB2 (PDB ID: 2RH1, 2R4R/2R4S [13, 14])[15]. It was solved using the LCP method that provides a more native, lipid environment for crystallization to a resolution of 2.4 Å. This delay was due to the need of numerous technological advancements required to crystallize membrane proteins like GPCRs. Developments in protein engineering, computational methods like homology

are asthma and strokes and cerebral hypoperfusion [10]. GPCRs control airway smooth muscle (ASM) contraction and increased airway resistance when coupled to Gq receptors. Airway epithelium and hematopoietic cells that are involved in control of lung inflammation that causes most asthma, have various pathways that are mediated by GPCRs. Arrestins regulate GPCR signaling and once again structural insights into the GPCRs is essential in understanding vital role of arrestins in those

GPCR-mediated airway cell functions that are dysregulated in asthma.

**3. A brief history on the structural study of GPCRs**

parts of functional importance in G-protein activation.

**58**

modeling and heterologous protein expressions have accelerated structural determination of GPCRs [16]. In this structure solved via the LCP technique, the proteins are placed in a membrane-like environment where they can diffuse and interact with each other to form crystal lattice contacts on both complementary hydrophobic and hydrophilic regions. These structures served as the template for the other crystal structures that were solved afterwards. Other receptors like H1R, D3R and 5-HT1B belonging to the Rhodopsin family of GPCRs were solved in the following years and served as templates for all the other GPCR structures that were predicted by homology modeling in the following years of research.

Another important subfamily of class A GPCRs with a number of key physiologic roles are the Chemokine receptors [17]. So far (till 2020) only 5 different chemokine receptor complexes have had their crystal structure solved by researchers and they are CXCR4 [18] (PDB IDs: 3ODU, 3OE0, 3OE6, 3OE8, 3OE9, and 4RWS [18, 19]), CCR5 [20] (PDB IDs: 4MBS, 5UIW, 6AKX, and 6AKY [21–23]), US28 [24] (PDB IDs: 4XT1, 4XT3, 5WB1, and 5WB2 [24, 25]), CCR2 [26] (PDB IDs: 5T1A, 6GPS, and 6GPX [26, 27]) and CCR9 [28] (PDB ID: 5LWE [28]). Structure based drug design was the key in solving crystal structures of Chemokine receptors and its potential is reflected by the large amount of ligands found for various chemokine receptors. SBDD methods prove to be more effective when a crystal structure is available as homology models.

In 2011, Kobilka achieved another break-through when he and his team captured an image of the β-adrenergic receptor at the exact moment that it is activated by a hormone and sends a signal into the cell. This image is a molecular masterpiece [29]. This was the first step in the path that earned Brian Kobilka the Nobel Prize in Chemistry in the year 2012 for his groundbreaking discoveries about GPCRs along with Robert Lefkowitz. In the year 2011 and 2013, the first secretin family GPCR structure was solved (PDB ID: 4L6R, 4K5Y [30, 31]) and in the following year the first glutamate family GPCR structure was deposited in PDB (PDB ID: 4OR2, 4OO9 [32, 33]).

There are various databases available exclusively for GPCR structures like GPCR-EXP [https://zhanglab.ccmb. med.umich.edu/GPCR-EXP/] (database for experimentally solved GPCR structures) and GPCRdb [34] (web tools and diagrams that aid GPCR research) that profusely help the researchers. According to GPCR-EXP statistics there are 389 structures for 67 GPCRs belonging to different species deposited in the PDB. **Figure 2** gives us details about the total number of new experimental

**Figure 2.** *The total number of new GPCR experimental structures available each year given by GPCR EXP database.*

structures of GPCRs solved every year as recorded by GPCR-EXP database. There are still many more GPCR structures that are yet to be solved and these remain as an unturned page in the global research of GPCRs.
