Page 66 - TD-3-1

Tumor Discovery Missense mutations in CXCR1: Impact on stability and function

1. Introduction magnetic resonance spectroscopy (NMR) investigations
have also reported that residues after the CXC sequence
CXCR1 is a rhodopsin-like (class A) G protein-coupled and those in the core β sheet of IL-8/CXCL8 bind directly
receptor (GPCR) that plays a crucial role in cellular signaling to the N-terminus of CXCR1. The ELR motif of the CXC
17
and is being investigated as a potential drug receptor due chemokine on the N-terminus of IL-8/CXCL8 plays a
to its involvement in various physiological processes. role in the interaction of the chemokine with CXCR1.
1,2
18
Rhodopsin is commonly used as a standard structure for Swapping the IC C-terminal regions consisting of 60 amino
mimicking GPCRs, and many rhodopsin structures have acids between CXCR1 and CXCR2 reportedly reversed
been characterized in protein databases (PDBs). CXCR1 the antagonistic effects of these receptors. Further
3
19
is one of two receptors that has a strong binding affinity studies have demonstrated that the relative antagonistic
for the CXC chemokine interleukin-8 (IL-8). IL-8 is also
4
known to regulate inflammatory and immune responses effects of CXCR1 on CXCR2 were determined by a single
amino acid residue: lysine 320 in CXCR2 and asparagine
and has been linked to various diseases, including cancer. 311 in CXCR1. The charged amino acids play a crucial
5
19
On exposure to inflammatory stimuli, the chemokine IL-8 role in maintaining the local structural stability required
binds to the extracellular (EC) region of CXCR1. Despite for binding in CXCR1, and mutating them disrupts this
6
the physiological importance of CXCR1, the underlying 6
molecular mechanism of its signal transduction pathway is stability and prevents binding. The mutations that were
poorly understood, primarily due to the limited availability studied involved charged amino acids in various regions of
20
of its structural information. 7 the CXCR1 receptor, including TM helices and EC loops.
These mutations were predicted to alter the local structure
GPCRs, including CXCR1, typically consist of seven of the receptor and potentially affect its ability to bind to
transmembrane (TM) helices, four EC regions (comprising other molecules. Specifically, the mutations were predicted
the N-terminal and three interhelical loops), and four to disrupt ion pairs, and oppositely charged amino acids
intracellular (IC) regions (including the C-terminal and would form bonds that can affect the receptor’s capacity to
three interhelical loops). Among the seven TM helices, maintain a stable structure in these regions. 21
8
TM3 and TM6 are regarded as pivotal elements responsible
for activating switches in GPCRs, and these regions play a Identifying deleterious mutations and predicting
critical role in the transition from a partially active state to a the effects of missense mutations on protein structure
fully activated state, particularly in the interactions between and function have become important applications of
22
G proteins and receptors. Crystal structures of the human computational analysis. This approach can aid in the
8,9
adenosine A2A receptor bound to rhodopsin and agonists design of new drugs and therapies for treating various
indicate that GPCR activation is induced by the disruption genetic diseases; for this reason, in the present study, we
of crucial interhelical interactions. Activation involves the evaluated the possible consequences of mutations in the
10
rotation of the TM3 and TM6 domains, thereby altering various CXCR1 domains that can affect protein function and
the conformational structure of the receptor’s G protein- interactions with ligands. To categorize harmful mutations,
interacting cytoplasmic loops to expose previously masked we initially used several computational algorithms,
24
25
23
G protein-binding sites on the IC loops. Recent advances including Meta-SNP, PMut, and PROVEAN. We also
11
in GPCR structural determination have studied the utilized the FATHMM server to detect specific genetic
molecular mechanisms of GPCR activation and activity in variants associated with carcinogenesis within disease-
27
26
various cellular processes, such as hormone signaling and causing mutations. Moreover, we employed mCSM,
28
29
12
neurotransmission. The techniques used to study GPCRs SDM, and DUET to predict the impact of these mutations
involve several modifications, including making changes on the structure and stability of the corresponding proteins.
to the receptors themselves, adding small molecules to The I-TASSER server was used to create a mutation model
stabilize them, and using specialized lipid mixtures to form with the necessary missense mutation, and Schrödinger’s
crystals that can be analyzed. 13,14 Activation of CXCR1 Desmond 3D modeling software was used to explore the
requires the involvement of both amino-terminal residues structural changes that affect the stability and function of
and EC loops. In this process, GPCR IC loops play a crucial the CXCR1 protein.
role in facilitating interactions with G proteins during 2. Methods
signaling. 4,15
2.1. Collection of datasets
According to experimental research, charge
significantly influences the interaction of CXCR1 with To obtain information on CXCR1 missense mutations, the
IL-8/CXCL8, and charged residues in the third and fourth study collected data from Ensembl (release 109) and the
30
EC loops are essential for this association. Nuclear Universal Protein Resource Knowledgebase (UniProtKB),
16
Volume 3 Issue 1 (2024) 2 https://doi.org/10.36922/td.2512