TRPM7 N-terminal region forms complexes with calcium binding proteins CaM and S100A1

Abstract

Transient receptor potential melastatin 7 (TRPM7) represents melastatin TRP channel with two significant functions, cation permeability and kinase activity. TRPM7 is widely expressed among tissues and is therefore involved in a variety of cellular functions representing mainly Mg 2þ homeostasis, cellular Ca 2þ flickering, and the regulation of DNA transcription by a cleaved kinase domain translocated to the nucleus. TRPM7 participates in several important biological processes in the nervous and cardiovascular systems. Together with the necessary function of the TRPM7 in these tissues and its recently analyzed overall structure, this channel requires further studies leading to the development of potential therapeutic targets. Here we present the first study investigating the N-termini of TRPM7 with binding regions for important intracellular modulators calmodulin (CaM) and calcium-binding protein S1 (S100A1) using in vitro and in silico approaches. Molecular simulations of the discovered complexes reveal their potential binding interfaces with common interaction patterns and the important role of basic residues present in the N-terminal binding region of TRPM.

TRPM5 Channel Binds Calcium-Binding Proteins Calmodulin and S100A1

“…39 The resulted complexes were compared with the TRPV1p/CaM and RyR1P12/S100A1 structures. 38,39 Consequently, we optimized the selected binding modes, as described by Bousova et al 35 The final TRPM5np/CaM and TRPM5np/S100A1 complexes confirmed the interactions of TRPM5np R85, R89, K90, and K94 with negatively charged residues of the ligands. TRPM5np interacted with these specific CaM negatively charged residues E11, E119, E123, and E127.…”

“…The interactions of TRP channels with CaM or S100A1 are profoundly maintained by TRP channels positively charged amino acid residues that form noncovalent interactions with CaM/S100A1 negatively charged amino acid residues. The clusters of positively charged residues at TRP channel binding epitopes commonly bear specific patterns in their positions. ,,, To predict the basic residues of TRPM5np involved in the CaM/S100A1complex interface, we analyzed the other TRPM binding region sequences for CaM (S100A1), which we had experimentally confirmed in our previous in vitro studies ,,, (Figure A, C). The selection of TRPM5np positively charged residues was performed by the sequence alignment of TRPM N-termini binding regions (Figure B).…”

“…The TRPM5np/CaM-Ca 2+ complex was built using ClusPro based on its similarity with the TRPV1p/CaM complex (PDB: 3SUI), as well as based on our previous docking TRPM/CaM strategy (Figure D, E). ,, The TRPM5np/S100A1-Ca 2+ complex was built using ClusPro based on the closest known S100A1/receptor-peptide structure of the RyR1P12/S100A1 (PDB: 2K2F) (Figure D, E) . The resulted complexes were compared with the TRPV1p/CaM and RyR1P12/S100A1 structures. , Consequently, we optimized the selected binding modes, as described by Bousova et al The final TRPM5np/CaM and TRPM5np/S100A1 complexes confirmed the interactions of TRPM5np R85, R89, K90, and K94 with negatively charged residues of the ligands. TRPM5np interacted with these specific CaM negatively charged residues E11, E119, E123, and E127.…”

“…Steady-state FA experiments were performed on a K2 spectrometer (ISS Inc., Champaign, Illinois, USA) at RT according to our previously described in-house FA protocol. 35 Lifetime Measurements. Fluorescence lifetimes were evaluated at RT using a confocal microscope IX83 (Olympus, Tokyo, Japan) according to our previously described in-house lifetime measurement protocol.…”

“…Fluorescence lifetimes were evaluated at RT using a confocal microscope IX83 (Olympus, Tokyo, Japan) according to our previously described in-house lifetime measurement protocol. 35 Microscale Thermophoresis. The CaM/S100A1 binding affinity to TRPM5np and TRPM5np_R85A/R89A/K90A/ K94A was characterized by MST using Monolith NT.115 (NanoTemper Technologies GmbH, Munich, Germany).…”