STUDY OF THE INTERACTION CAPACITY OF PSEUDO-PHOSPHORYLATED RECOMBINANT PEPTIDES WITH PLCg

• Grant number: 25/05336-4
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: July 01, 2025
• End date: June 30, 2026
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Deborah Schechtman
• Grantee: Eloísa Hermann Moser
• Host Institution: Instituto de Química (IQ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 19/06982-6 - Characterization and development of new modulators of the TrkA and PKMzeta pathways in inflammatory and chronic pain, AP.TEM

Abstract

Even though it is an alert of utmost importance for an organism's survival, pain can lead to anguish that requires the use of medication to contain it. Nowadays, the available analgesics do not present the desired efficacy, especially for chronic pain, and cause side effects that affect users in many spheres, underscoring the necessity for developing new strategies to manage pain. In this regard, the study of intracellular signalling pathways leading to pain is highlighted as a target for obtaining information that will allow the development of new analgesics. Amongst these pathways, the one related to phospholipase C gamma (PLCg), activated through the interaction between nerve growth factor (NGF) and tropomyosin receptor kinase A (TrkA), distinguishes itself as relevant in pain signalling. After tissue injury, NGF secreted by inflammatory cells binds to its receptor tyrosine kinase (TrkA) present in nociceptor neurons, promoting autophosphorylation of the receptor. In this context, the phosphorylated tyrosines within the cytoplasmic domains of TrkA act as anchoring sites for substrates, such as PLCg, which is then phosphorylated by the receptor's kinase activity, thereafter inducing a series of steps that result in activating the ion channel regarded as transient receptor potential cation channel subfamily V member 1 (TRPV1), which leads to the propagation of pain through the nervous system. Given this information, our laboratory developed a peptide based on the anchoring site of PLC¿¿ on TrkA, called TAT-pQYP, which binds to the N-SH2 domain of the phospholipase preventing its binding and subsequent phosphorylation/activation by the receptor, inhibiting PLCg-mediated signalling and, hence, reducing mechanical sensitivity in animal assays. In sight of the inhibiting activity showcased by the aforementioned peptide, this project aims to analyze, through polarization assays, the affinity of other peptides, derived from TAT-pQYP, to PLCg, as well as the feasibility of producing such peptides through heterologous expression. Peptides derived from TAT-pQYP will be developed by substituting the phosphorylated tyrosine residue for a glutamic acid residue or a phenylalanine residue, thus being named QEP and QFP, respectively. Therefore, the viability of using peptides that are less complex and expensive in studies related to pain therapeutics will be verified as well as the inhibiting potentialities of smaller peptides with phosphomimetic residues, so that these can possibly assist in conceptualizing more efficient analgesics. (AU)