MOLECULAR SWITCHES

Seminar Bioorganik (WS18/19)

Leipzig, 28.01.2019

INTRODUCTION

Molecular switches

OPTOGENETICS

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= technique to control cells in living tissue (typically

neurons) by light

Cells were genetically modified to express light-sensitive

ion channels

Optochemical genetics → addition of a chemical

component

Molecular switches

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transmembrane proteins, sensitive to an input signal:

Light intensity

Pressure

Voltage

Temperature

Ligand concentration

all can be influenced by ligands (e.g. channel blockers)

→ point of contact for optochemical genetics

TRANSMEMBRANE RECEPTORS IN

NEUROBIOLOGY

Molecular switches

TRANSMEMBRANE RECEPTORS IN NEUROBIOLOGY

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Activated by / ligand / voltage

Molecular switches

NEUROTRANSMISSION

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Molecular switches

CHANNELRHODOPSIN-2

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Molecular switches

LIGHTING UP THE BRAIN – LIGHT-SENSITIVE

RECEPTORS

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→ Optogenetics

Fast modulation of intensity within femtoseconds

Can be focused onto very small areas

Carry enough energy to trigger larger molecular motions

MOLECULAR TOOLS

IN OPTOCHEMICAL

GENETICS

Molecular switches

CAGED LIGAND APPROACH (CL)

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Simplest and oldest approach

first shown in living animals

Irreversible process and produces by-products

Molecular switches

PHOTOCHROMIC LIGAND APPROACH (PCL)

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Ease of application, fast distribution → good as drugs

Work well in complex systems

Selectivity between receptor subtypes is challenging

Molecular switches

PHOTOSWITCHED TETHERED LIGAND

APPROACH (PTL)

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Selectivity between receptor subtypes

Can be genetically encoded → biologic, not synthetic

Ligand does not need high affinity

Molecular switches

PTL – FUNKTIONAL DISSECTION OF RELATED

RECEPTOR SUBTYPES

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Molecular switches

AZOBENZENE PHOTOSWITCHES

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Fast, photostable, need low intensity

Easy to synthesize and modify

Low solubility → overcome with functional groups

VOLTAGE-GATED

POTASSIUM

CHANNELS

Molecular switches

VOLTAGE-GATED POTASSIUM CHANNELS

Modulate cellular excitability

Play a key role in the generation of action potentials

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Molecular switches

VOLTAGE-GATED POTASSIUM CHANNELS

Channel blockers

Cs+ ions

Venom peptides

Small organic cations

External TEA

Internal TEA

17TEA: tetraethylammonium

Molecular switches

PHOTOSENSITIZING VOLTAGE-GATED

POTASSIUM CHANNELS

PTL approach

H-SPARK

Hyperpolarizing effect

18SPARK: Synthetic photoisomerizable azobenzene-regulated K+ channel

MAQ: maleimide/azobenzene/quartenary/ammonium

Molecular switches

PHOTOSENSITIZING VOLTAGE-GATED

POTASSIUM CHANNELS

PTL approach

D-SPARK

Depolarizing effect

19SPARK: Synthetic photoisomerizable azobenzene-regulated K+ channel

Molecular switches

PHOTOSENSITIZING VOLTAGE-GATED

POTASSIUM CHANNELS

PCL approach

XAQs

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Molecular switches

PHOTOSENSITIZING VOLTAGE-GATED

POTASSIUM CHANNELS

Shift the absorption and action spectra toward the red

deeper tissue penetration

diminish the phototoxity

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IONOTROPIC

GLUTAMATE

RECEPTORS

Molecular switches

IONOTROPIC GLUTAMATE RECEPTORS

Play a cenral role in synaptic transmission

Antagonists

„foot-in-the-door“ mechanism

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Molecular switches

PHOTOSENSITIZING IONOTROPIC GLUTAMATE

RECEPTORS

PTL approach

LiGluR

24LiGluR: light-gated ionotropic receptor

Molecular switches

PHOTOSENSITIZING IONOTROPIC GLUTAMATE

RECEPTORS

Sign inversion

MAG-0 is attached to

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cis activation

trans activation

Molecular switches

PHOTOSENSITIZING IONOTROPIC GLUTAMATE

RECEPTORS

HyLighter

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K+ selective TMD of the

procaroytic glutamate

receptor

Photosensitive LBD of

LiGluR

Molecular switches

PHOTOSENSITIZING IONOTROPIC GLUTAMATE

RECEPTORS

Controlling the escape reflex in zebrafish

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Molecular switches

PHOTOSENSITIZING IONOTROPIC GLUTAMATE

RECEPTORS

PCL approach

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PENTAMERIC LIGAND-

GATED ION CHANNELS

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Molecular switches

SUBGROUPS

Nicotinic acetylcholine receptor (nAChR)

γ-aminobutyric acid receptors (GABAA/C)

5-hydroxytryptamine receptors (5-HT3)

glycine receptors (GlyRs)

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acetylcholine

GABA

serotonin

glycine

Molecular switches

NICOTINIC ACETYLCHOLINE RECEPTOR

31Source: Nasiripourdori, A.; et al.: From Toxins Targeting Ligand Gated Ion

Channels to Therapeutic Molecules. Toxins. December 2011, 3:260-293

Molecular switches

Source:

NICOTINIC ACETYLCHOLINE RECEPTOR

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• first photosensitized

receptor

• easy accessable in

certain fishes

acetylcholine

or

nicotin

Electrophoruselectricus

Source: https://de.wikipedia.org/wiki/Zitteraal#/media/File:Electrophorus_electricus_3.jpg,

18.01.19

Molecular switches

PHOTOSENSITIZING OF NICOTINIC

ACETYLCHOLINE RECEPTOR

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PCL

PTL

OUTLOOK

Molecular switches

ALTERNATIVE AND FUTURE TARGETS FOR

PHOTOSENSITIZING

• countless receptors

• taste and olfactory receptors, GPCRs, etc.

• receptor linked enzymes

• e. g. receptor tyrosine kinases

• receptor agonists and antagonists

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Molecular switches

PHOTOACTIVATABLE CHEMOKINES

36Source: Baumann, L.; Beck-Sickinger, A. G.: Photoactivatable Chemokines – Controlling Protein

Activity by Light. Angewandte Chemie. June 2013, Volume 52, Issue 36

Molecular switches

FUTURE TASKS AND UTILISATIONS

• phoswitchable cross-linkers

• exploring of new photoswitchers

• switchers sensitive für other signals

• orthogonal ligands

• electromagnetic fields

• knock-in instead of knock-out

mutants

37Source: Valeur, E.; et al.: New Modalities for Challenging Targets in Drug Discovery. Angewandte

Chemie. February 2017, Volume 56, Issue 35

Molecular switches

CURING BLINDNESS

38Source: https://www.allaboutvision.com/resources/retina.htm (20.01.19)

THANK YOU FOR YOUR

ATTENTION!