Williams BCH 444 2014 Lectures 3-6

Alu domain s domain srp54 m domain is helical with a

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Unformatted text preview: recognize so many different signal sequences? Alu domain S domain - SRP54 M domain is α-helical with a hydrophobic groove formed by helices αM1, αM2, αM4 and a finger loop between αM1 and αM2. - Content of Met residues in SRP54 is 16% - six times higher than average - Met sidechain is hydrophobic and, due to unbranched nature and thioether linkage, conformationally flexible. -  Thought that this hydrophobic groove lined by many methionine-bristles allows SRP sufficient plasticity to recognize diverse hydrophobic signal squences. 7S RNA in red + yellow hydrophobic groove SRP54 M domain α-helical signal sequence (modeled) likely that RNA backbone recognizes n- or c-region of signal sequence 7SRNA SRP54 M domain Keenan, R.J. et al (2001) Ann. Rev. Biochem. 70:755 14 Bringing the ribosome to the ER - the SRP-SRP receptor interaction 1. Proteins destined for the ER are synthesized (usually) with an N-terminal signal sequence 2. Signal is recognized by Signal Recognition Particle (SRP) 3. SRP binds to SRP receptor on ER membrane - (brings translating polyribosome to the ER). SRP receptor helps target the ribosome to the translocon pore. 4.  SRP is released, ribosome binds to translocon, nascent chain enters translocon. 5. Nascent polypeptide is translocated into the ER lumen and signal is removed by signal peptidase 1 2 4 3 5 15 The SRP-SRP receptor interaction - a tale of three GTPases ribosome stimulates GTP binding to SRP54 but signal sequence inhibits hydrolysis 1.  SRP receptor (SR) consists of two subunits: SRα, a soluble GTPase, and SRβ, a membrane-bound GTPase 2.  SRP54 not only binds signal sequences through its M domain but its NG domain is a GTPase that binds to the GTPase domain of SR receptor α subunit. 3. GTP binding is required for SRP-SR interaction and hydrolysis causes dissociation. Note: GTP hydrolysis is stimulated upon SRP-SR binding, i.e., SRP acts as a GTPase-activating protein (GAP) for SR and vice versa. 4. SRβ must be in GTP form to bind to SRα. SRβ seems to be the SR subunit that interacts with the translocon pore. SRP and SR act as GTP-regulated molecular matchmakers. SRP picks up ribosomes with exposed signal sequences in the cytosol and SR picks up empty translocons in the ER membrane. End result is targeting of nascent chains to the translocon. Egea, PF et al. (2005) Curr. Opin. Struct. Biol. 15:213 Interaction between SRP54 GTPase domain and GTPase domain of SRα (shared GTPase active site) GTP-dependent interaction between SRβ and non-GTPase X-domain of SRα 16 Translocation of nascent polypeptide across the ER membrane 1. Proteins destined for the ER are synthesized (usually) with an N-terminal signal sequence 2. Signal is recognized by Signal Recognition Particle (SRP) 3. SRP binds to SRP receptor on ER membrane - (brings translating polyribosome to the ER). SRP receptor helps target the ribosome to the translocon pore. 4.  SRP is released, ribosome binds to translocon, nascent chain enters translocon. 5. Nascent polypeptide is translocated into the ER lumen and signal is removed by signal peptidase Translocation 1 2 4 3 5 17 First Quiz kDa Stds -mics -mics + protease 1 2 +mics +mics + protease 100 70 45 20 3 4 18 BCH 444 Spring 2014 •  Targeting nascent proteins to the endoplasmic reticulum •  Protein translocation into the ER, membrane protein topology •  Protein glycosylation, functions of glycoproteins •  Protein folding in the ER, role of chaperones and folding cat...
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This note was uploaded on 03/27/2014 for the course BCH 444 taught by Professor Mccallan during the Spring '14 term at University of Toronto- Toronto.

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