Recombinant mouse Mast cell protease 4 (Mcpt4) production begins with gene cloning. The gene encoding the Mcpt4 protein (21-246aa) is inserted into a plasmid vector along with the N-terminal 6xHis-tag gene and introduced into yeast cells. The cells are cultured in bioreactors to produce the Mcpt4 protein. Once sufficient protein is produced, the cells are lysed, and the Mcpt4 protein is purified through affinity chromatography. The final product is subjected to SDS-PAGE to confirm its purity. Its purity is greater than 90%.
Mcpt4 is a chymase enzyme found in mice that plays a crucial role in various physiological processes. In mice, Mcpt4 is considered the functional homolog to human chymase [1]. Studies have shown that Mcpt4 has pro- and anti-inflammatory roles depending on the disease model [2]. Additionally, Mcpt4 has been implicated in processes such as bone density regulation [3], recruitment of leukocytes in inflammation [2], degradation of insulin-like growth factor-1 leading to adverse cardiac remodeling [4], suppression of scar formation after spinal cord injury [5], and protection against melanoma colonization of the lung [6].
Furthermore, Mcpt4 has been associated with protective roles in host resistance to venom toxicity [7], regulation of thrombin and fibronectin turnover [8], and suppression of the host immune response to malaria [9]. Studies have also highlighted the involvement of Mcpt4 in wound healing processes [10], itch induced by endothelin-1 [11], and regulation of coagulation factor XIIIA levels.
References:
[1] T. Lind, A. Gustafson, G. Calounova, L. Hu, A. Rasmusson, K. Jonssonet al., Increased bone mass in female mice lacking mast cell chymase, Plos One, vol. 11, no. 12, p. e0167964, 2016. https://doi.org/10.1371/journal.pone.0167964
[2] J. Succar, G. Giatsidis, N. Yu, K. Hassan, R. Khouri, M. Gurishet al., Mouse mast cell protease-4 recruits leukocytes in the inflammatory phase of surgically wounded skin, Advances in Wound Care, vol. 8, no. 10, p. 469-475, 2019. https://doi.org/10.1089/wound.2018.0898
[3] T. Tejada, L. Tan, R. Torres, J. Calvert, J. Lambert, M. Zaidiet al., Igf-1 degradation by mouse mast cell protease 4 promotes cell death and adverse cardiac remodeling days after a myocardial infarction, Proceedings of the National Academy of Sciences, vol. 113, no. 25, p. 6949-6954, 2016. https://doi.org/10.1073/pnas.1603127113
[4] T. Vangansewinkel, S. Lemmens, N. Geurts, K. Quanten, D. Dooley, G. Pejleret al., Mouse mast cell protease 4 suppresses scar formation after traumatic spinal cord injury, Scientific Reports, vol. 9, no. 1, 2019. https://doi.org/10.1038/s41598-019-39551-1
[5] M. Grujić, A. Paivandy, A. Gustafson, A. Thomsen, H. Öhrvik, & G. Pejler, The combined action of mast cell chymase, tryptase and carboxypeptidase a3 protects against melanoma colonization of the lung, Oncotarget, vol. 8, no. 15, p. 25066-25079, 2017. https://doi.org/10.18632/oncotarget.15339
[6] M. Akahoshi, C. Song, A. Piliponsky, M. Metz, A. Guzzetta, M. Åbrinket al., Mast cell chymase reduces the toxicity of gila monster venom, scorpion venom, and vasoactive intestinal polypeptide in mice, Journal of Clinical Investigation, vol. 121, no. 10, p. 4180-4191, 2011. https://doi.org/10.1172/jci46139
[7] E. Tchougounova, G. Pejler, & M. Åbrink, The chymase, mouse mast cell protease 4, constitutes the major chymotrypsin-like activity in peritoneum and ear tissue. a role for mouse mast cell protease 4 in thrombin regulation and fibronectin turnover, The Journal of Experimental Medicine, vol. 198, no. 3, p. 423-431, 2003. https://doi.org/10.1084/jem.20030671
[8] N. Céspedes, E. Donnelly, C. Lowder, G. Hansten, D. Wagers, A. Briggset al., Mast cell chymase/mcpt4 suppresses the host immune response to plasmodium yoelii, limits malaria-associated disruption of intestinal barrier integrity and reduces parasite transmission to anopheles stephensi, Frontiers in Immunology, vol. 13, 2022. https://doi.org/10.3389/fimmu.2022.801120
[9] J. Succar, J. Douaiher, L. Lancerotto, Q. Li, R. Yamaguchi, G. Younanet al., The role of mouse mast cell proteases in the proliferative phase of wound healing in microdeformational wound therapy, Plastic & Reconstructive Surgery, vol. 134, no. 3, p. 459-467, 2014. https://doi.org/10.1097/prs.0000000000000432
[10] E. Magnúsdóttir, M. Grujic, J. Bergman, G. Pejler, & M. Lagerström, Mouse connective tissue mast cell proteases tryptase and carboxypeptidase a3 play protective roles in itch induced by endothelin-1, Journal of Neuroinflammation, vol. 17, no. 1, 2020. https://doi.org/10.1186/s12974-020-01795-4
[11] N. Shubin, V. Glukhova, M. Clauson, P. Truong, M. Åbrink, G. Pejleret al., Proteome analysis of mast cell releasates reveals a role for chymase in the regulation of coagulation factor xiiia levels via proteolytic degradation, Journal of Allergy and Clinical Immunology, vol. 139, no. 1, p. 323-334, 2017. https://doi.org/10.1016/j.jaci.2016.03.051
