Ala5-galanin (2-11) is a GAL2R specific galanin analogue.
It is over 30years since the regulatory peptide galanin was discovered by Professor Mutt and co-workers. Galanin exerts its effects by binding to three galanin G-protein coupled receptors, namely GAL1R, GAL2R and GAL3R. Each galanin receptor has a different distribution in the central nervous system and the peripheral nervous system as well as distinctive signaling pathways, which implicates that the receptors are involved in different biological- and pathological effects. The delineation of the galaninergic system is however difficult due to a lack of stable, specific galanin receptor ligands. Herein, a new short GAL2R specific ligand, Ala5-galanin (2-11), is presented. The galanin (2-11) modified analogue Ala5-galanin (2-11) was tested in125I-galanin competitive binding studies for the three galanin receptors and the G-protein coupled receptor signaling properties was tested by the ability to influence second-messenger molecules like inositol phosphate and cyclic adenosine monophosphate. In addition, two different label-free real-time assays, namely EnSpire® based on an optical biosensor and xCELLigence® based on an electric biosensor, were used for evaluating the signaling properties using cell lines with different levels of receptor expression. Ala5-galanin (2-11) was subsequently found to be a full agonist for GAL2R with more than 375-fold preference for GAL2R compared to both GAL1R and GAL3R. The single amino acid substitution of serine to alanine at position 5 in the short ligand galanin (2-11) resulted in a ligand subsequently unable to bind neither GAL3R nor GAL1R, even at concentrations as high as 0.1mM.
Department of Neurochemistry, Stockholm University, Svante Arrheniusv. 16B, SE-10691 Stockholm, Sweden. Electronic address: kristin.webling@neurochem.su.se.
Kristin
K
Webling
Department of Neurochemistry, Stockholm University, Svante Arrheniusv. 16B, SE-10691 Stockholm, Sweden.
Johan
J
Runesson
Research Program for Receptorbiochemistry and Tumormetabolism, Laura Bassi Centre of Expertise THERAPEP, Department of Pediatrics, University Hospital Salzburg, Paracelsus Medical University, Müllner Hauptstr. 48, A-5020 Salzburg, Austria.
Andreas
A
Lang
Institute of Technology, University of Tartu, Nooruse 1, 50411 Tartu, Estonia.
Indrek
I
Saar
Research Program for Receptorbiochemistry and Tumormetabolism, Laura Bassi Centre of Expertise THERAPEP, Department of Pediatrics, University Hospital Salzburg, Paracelsus Medical University, Müllner Hauptstr. 48, A-5020 Salzburg, Austria.
Barbara
B
Kofler
Department of Neurochemistry, Stockholm University, Svante Arrheniusv. 16B, SE-10691 Stockholm, Sweden; Institute of Technology, University of Tartu, Nooruse 1, 50411 Tartu, Estonia.
Ülo
Ü
Langel
D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D002460:Cell Line; D003412:Cricetulus; D019004:Galanin / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D044092:Receptor, Galanin, Type 2 / Q000819:agonists* / Q000378:metabolism
D010446:Peptide Fragments; D044092:Receptor, Galanin, Type 2; D019004:Galanin
Agonist; Dynamic mass redistribution; GAL(2)R specific ligand; GPCR; Galanin; Impedance; Label-free real-time assay; Neuropeptide