Publications
Application notes and peer-reviewed publications
Application Notes
Salipro® direct membrane extraction
Salipro® goes DirectMX™
Here, we present DirectMX™ for the direct extraction of membrane proteins from crude cell membranes into Salipro® particles with native lipids. The method presents membrane proteins in Salipro® which can be treated like soluble proteins and are embedded in a detergent-free, natural lipid environment. This opens up new opportunities for the analysis of membrane proteins and the identification of novel drug targets.
Salipro® for drug screening
Salipro® goes Biacore™
Structural and functional studies of membrane proteins as part of drug discovery efforts are limited by their poor stability outside the native membrane environment. This represents a major challenge for the pharmaceutical industry. Here, in collaboration with GE Healthcare and EMBL Hamburg, we utilized the Salipro® lipid nanoparticle system for binding studies of membrane proteins using Biacore™ systems, in a detergent-free environment.
Salipro® for antibody discovery
Salipro® goes immunization and B cell sorting
The key to making functional monoclonal antibodies (mAbs) is to use pure, stable, homogeneous antigen for immunization, mAb selection/sorting and for in vitro HTS characterization. The Salipro® technology enables all of the above for fragile membrane protein targets. In addition, Salipro® represents a validated approach for membrane protein epitope mapping by cryo-EM.
Salipro® for antibody discovery
Salipro® for cryoEM
GEN Drug Discovery Tutorial:
Salipro® goes phage display
To date there is no technology platform that has been able to routinely generate therapeutic leads directed at membrane protein targets. Here we describe how the Salipro® platform technology enables phage display selection and downstream antibody characterization with a purified ion channel.
Salipro® goes cryoEM
Cryo-electron microscopy (cryo-EM) is rapidly evolving to be the primary tool for the structure determination of membrane proteins. We asked whether it would be possible to generate high-quality pannexin 1 ion channel protein with the Salipro platform for structure elucidation by cryo-EM. Here, we present the three stages of our approach – protein expression, generation of purified Salipro®- pannexin 1, and structure determination.
Publications
S. Basse Hansen, et al., Structure of a Ca2+ bound phosphoenzyme intermediate in the inward-to-outward transition of Ca2+-ATPase 1 from Listeria monocytogenes. bioRxiv doi: 10.1101/2024.03.06.583647 (2024).
J. Zhang, et al., Open structure and gating of the Arabidopsis mechanosensitive ion channel MSL10. Nat. Commun. 14, 6284 (2023).
J. Cowgill, et al., Structure and dynamics of differential ligand binding in the human p-type GABAA receptor. Neuron 111, 1–15 (2023).
D. H. Legesse, et al., Structural insights into opposing actions of neurosteroids on GABAA receptors. Nat. Commun. 14, 5091(2023).
U. Goswami, et al., Structural interplay of anesthetics and paralytics on muscle nicotinic receptors. Nat. Commun. 14, 3169 (2023).
I. Drulyte, et al., Direct cell extraction of membrane proteins for structure-function analysis. Scientific Reports 13, 13:1420 (2023). (Describes Salipro® for cryoEM and SPR)
B. Xu, et al., Embigin facilitates monocarboxylate transporter 1 localization to the plasma membrane and transition to a decoupling state. Cell Reports 40, 111343 (2022).
S. Zhu, et al., Structural and dynamic mechanisms of GABAAreceptor modulators with opposing activities. Nat. Commun. 13, 4582 (2022).
K. Zhang, et al., Fusion protein strategies for cryo-EM study of G protein-coupled receptors. Nat. Commun. 13, 4366 (2022).
C. M. Noviello, et al., Structural mechanisms of GABAA receptor autoimmune encephalitis. Cell 185, 2469-2477 (2022).
M. M. Rahman, et al., Structural mechanism of muscle nicotinic receptor desensitization and block by curare. Nat. Struct. Mol. Biol. 29, 386-394 (2022).
C. Wang, M. M. Polovitskaya, B. D. Delgado, T. J. Jentsch, S. B. Long, Gating choreography and mechanism of the human proton-activated chloride channel ASOR. Sci. Adv. 8, 1–13 (2022).
M. M. Rahman, B. T. Worrell, M. H. B. Stowell, R. E. Hibbs, Purification of a native nicotinic receptor. Methods Enzymol. 653, 189–206 (2021).
D.-M. Kehlenbeck, et al., Cryo-EM structure of MsbA in saposin-lipid nanoparticles ( Salipro ) provides insights into nucleotide coordination. FEBS J., 1–12 (2021).
A. Ornik-cha, et al., Structural and functional analysis of the promiscuous AcrB and AdeB efflux pumps suggests different drug binding mechanisms. Nat. Commun. 12, 6919 (2021).
R. J. Howard, Elephants in the Dark: Insights and Incongruities in Pentameric Ligand-gated Ion Channel Models. J. Mol. Biol. 433, 167128 (2021).
F. Zhou, et al., Footprinting Mass Spectrometry of Membrane Proteins: Ferroportin Reconstituted in Saposin A Picodiscs. Anal. Chem. (2021).
L. Xiao, et al., Structures of the β-barrel assembly machine recognizing outer membrane protein substrates. FASEB J. 35, 1–13 (2021).
C. M. Noviello, et al., Structure and gating mechanism of the a7 nicotinic acetylcholine receptor. Cell 184, 1–14 (2021).
Y. Zhang, et al., Asymmetric opening of the homopentameric 5-HT3A serotonin receptor in lipid bilayers. Nat. Commun. 12, 1074 (2021).
J. J. Kim, et al., Shared structural mechanisms of general anaesthetics and benzodiazepines. Nature (2020).
T. Lasitza-Male, et al., Membrane Chemistry Tunes the Structure of a Peptide Transporter. Angew. Chemie Int. Ed. (2020).
P. Lloris-Garcerá, et al., DirectMX – One-Step Reconstitution of Membrane Proteins From Crude Cell Membranes Into Salipro Nanoparticles. Front. Bioeng. Biotechnol. 8, 1–9 (2020). (Describes Salipro® DirectMX™)
D. Du, et al., Interactions of a bacterial RND transporter with a transmembrane small protein in a lipid environment. Struct. Des., 1–10 (2020).
M. M. Rahman, et al., Structure of the Native Muscle-type Nicotinic Receptor and Inhibition by Snake Venom Toxins. Neuron, 1–11 (2020).
A. Gharpure, et al., Agonist Selectivity and Ion Permeation in the α3β4 Ganglionic Nicotinic Receptor. Neuron 104, 501-511.e6 (2019).
R. Nagamura, et al., Structural basis for oligomerization of the prokaryotic peptide transporter PepT So2. Acta Crystallogr. Sect. F Struct. Biol. Commun. 75, 348–358 (2019).
D. M. Kehlenbeck, et al., Comparison of lipidic carrier systems for integral membrane proteins - MsbA as case study. Biol. Chem. (2019).
K. Kanonenberg, S. H. J. Smits, L. Schmitt, Functional Reconstitution of HlyB, a Type I Secretion ABC Transporter, in Saposin-A Nanoparticles. Sci. Rep. 9, 1–12 (2019).
N. X. Nguyen, et al., Cryo-EM structure of a fungal mitochondrial calcium uniporter. Nature 559, 570–574 (2018).
A. Flayhan, et al., Saposin Lipid Nanoparticles: A Highly Versatile and Modular Tool for Membrane Protein Research. Structure 26, 345-355.e5 (2018).
A. F. Kintzer, et al., Structural basis for activation of voltage sensor domains in an ion channel TPC1. Proc. Natl. Acad. Sci. U. S. A. 115, E9095–E9104 (2018).
J. A. Lyons, A. Bøggild, P. Nissen, J. Frauenfeld, Saposin-Lipoprotein Scaffolds for Structure Determination of Membrane Transporters. Methods Enzymol. 594, 85-99 (2017).
C. T. H. Chien, et al., An Adaptable Phospholipid Membrane Mimetic System for Solution NMR Studies of Membrane Proteins. J. Am. Chem. Soc. 139, 14829–14832 (2017).
J. Frauenfeld, et al., A saposin-lipoprotein nanoparticle system for membrane proteins. Nat. Methods 13, 345–351 (2016). (Introduction to Salipro®)