G Proteins- Part 2

Biochemistry 4000

Dr. Ute Kothe

Ras-activated MAP kinase

cascade

Voet, Biochemistry

Heterotrimeric G proteins

Voet, Biochemistry

90º

• three subunits: , associate only in GDP form• subunit = GTPase• & subunits are linked to membrane by covalently attached lipids• become activated by membrane receptor (GEF)• become inactivated by “special GAPs” called Regulators of G protein signaling (RGS)

G proteins

Voet, Biochemistry

• at least 27 different Gproteins in humans

• three Switch regions

• extra helical domain inserted into canonical G domain

Gt (transducin) – involved in vision

G protein coupled Receptors

Voet, Biochemistry

G protein coupled receptors (GPCR):

• 7 transmembrane receptors

• GEF for heterotrimeric G proteins

• activation of G induces dissociation of G

• large class of receptors, e.g. for epinephrine (adrenergic receptors), odorant molecules, light (rhodopsin)

Activation & Inhibition of Adenyl Cyclase (AC) by G proteins

Voet, Biochemistry

Gi1-RGS4 complex

Tesmer, Cell 1997

• Gi1-Mg2+-GDP-AlF4- -RGS4

• 2.8 Å, molecular replacement using Gi1-GDP-AlF4- structure

• Rcryst = 29%, Rfree = 21%

RGS4 contacts all switch regions of Gi1 with the bottom of its four-helix bundle

Interface between RGS4 & Gi1

Tesmer, Cell 1997

RGS4 loop 3-4, loop 5-6 and loop 7-8 interact with all three switch regions of Gi1

RGS4 reduces the flexibility of all three switch regions (comparison of B factors of this structure with Gi1 alone)

Active Site of Gi1

Tesmer, Cell 1997

• no catalytic residue added by RGS4

• GTPase activation by stabilization of the switch regions, i.e. stabilization of the transition state

• RGS4-Asn128 might facilitate GTP hydrolysis either by stabilizing Gln 204 or by orienting the attacking H2O

• catalytic Arginine 178 provided in cis

Visual preception

Lodish, 5th edition

Rhodopsin is activated by light

Acts as GEF for transducin (G only found in rod cells)

Phototransduction cascade

Lodish, 5th edition

Rhodopsin Transducin (Gta) cGMP phosphodiesterase (PDE) cGMP GMP cGMP dissociates from nucleotide-gated channel Channel closes Hyperpolarization of membrane

Light Detection by Retinal

Lodish, 5th edition

Retinal is covalently bound to Rhodopsin

Isomerization of Retinal results in conformational change of rhodopsin (activation)

Organization of the olfactory system

Olfactory cortex

Main olfactory bulb

Sensory neurons in Olfactory ephithelium

Nasal cavity

Olfactory signal transductionOdorant Receptors:

• discovered in 1991

Nobel prize for Linda Buck & Richard Axel 2004

• GPCRs

• largest multigene family

• ~ 350 in human

• ~ 1000 in mice (1-3 % of the genes)