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Original Research Article
Screening and Mechanism of
Antagonist Peptide for CC Chemokine Receptor 1 (CCR1)
Derived from Viral Macrophage Inflammatory Protein II
Sha Liu, Qing Ding, Pijin
Wei, Hanxiao Sun*, Xiuying Li, Guijie An, Yan Yang and
Jingguang Zhou
Institute of Genomic
Medicine, College of Pharmacy, Jinan University,
Guangzhou 510632, China
*For correspondence:
Email:
sunhx718@163.com; Tel:
+86-020-38375022
Received: 11 February 2014
Revised accepted: 20 April 2014
Tropical
Journal of Pharmaceutical Research, May 2014;
13(5): 697-704
http://dx.doi.org/10.4314/tjpr.v13i5.7
Abstract
Purpose: To search for effective
antagonist peptide of CC chemokine receptor
1 (CCR1), and evaluate the potential role
and mechanism of peptide C18P derived from
viral macrophage inflammatory protein II (vMIP-II).
Methods: Alignment, simulated
peptide-cut, bioinformatics and protease
digestion were used to screen and prepare
antagonist peptide. Interactions between
C18P and CCR1 were determined by radioligand
binding assays and [35S]GTPγS binding
experiment. Chemotaxis assay was utilized to
assess the potency for inducing or
inhibiting peripheral blood mononuclear
cells (PBMCs) migration. Ligand-induced
intracellular calcium mobilization was
tested by flow cytometry.
Results: A peptide containing 18 amino acids
(C18P) was screened. C18P bound to CCR1 with
a Kd of 5.7 ng/ml and displaced 125I-labeled
MIP-1α and 125I-labeled RANTES on human
CCR1-transfected HEK293 cells (IC50 = 11.2
and 9.6 ng/ml, respectively) in radioligand
binding studies. C18P lacked intrinsic
agonist activity but strongly inhibited
HCC-1 (100 nM) induced [35S]GTPγS binding
(IC50 = 3.7 ug/ml), chemotaxis induced by
HCC-1, MIP-1α and RANTES (IC50 = 23, 25 and
13.1 ng/ml, respectively), and intracellular
calcium mobilization.
Conclusion: These results demonstrate
that bioinformatics and protease digestion
are feasible to screen and prepare C18P, and
that C18P is a novel and specific small
molecule peptide antagonist of CCR1 with
therapeutic potential for preventing cell
migration.
Keywords: CC Chemokine receptor 1;
Simulated peptide-cut; Antagonist peptide;
Viral macrophage inflammatory protein II;
Bioinformatics; Protease digestion; HEK293
cells; Radioligand binding
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