Download Protein folding and movement in the bacterial cell The action of

Protein folding and movement
in the bacterial cell
• All protein synthesis occurs in cytoplasm
• Generally, product of translation is unfolded
polypeptide, which must fold into proper 3
dimensional structure in order to function
! Polypeptide folding often will start before
translation is finished, with " helices & # strands (Fig.
3.15) forming spontaneously
!Tertiary/Quaternary (3°/4°) protein folding can
occur spontaneously but frequently is aided by
molecular chaperones
• At least 20% of all polypeptides made ultimately are
localized outside of the cytoplasm
Localization of proteins to
different cellular compartments
After synthesis in cytoplasm, proteins destined for
different cellular compartments are targeted there in
different ways:
• “Secreted proteins” that leave the cytoplasm before
3°/4° folding occurs, using a targeting signal that is
removed during export
• Proteins that are inserted into the cytoplasmic
membrane (where they will fold)
• Folded proteins that are exported with co-factors
The action of chaperones
Chaperones often
hydrolyze ATP
during 3°/4° folding
of cytoplasmic
proteins
Unfolded or
Chaperones can help
protein attain its
structure, but are
not part of the final
structure
Hsp70
Hsp60
Fig. 7.32
Targeting signals for protein export across
cytoplasmic membrane
Proteins destined to cross the cytoplasmic
membrane for final localization outside the
cell (or in the periplasm/outer membrane of
Gram neg. bacteria) generally have an Nterminal sequence that directs polypeptide to
machinery that carries out the localization.
One class of these targeting signals are used in
both proks and euks to direct precursor
proteins for secretion: signal sequences (SS
or signal peptides or leader sequences).
These SS are cleaved from precursor during
localization process.
General features of Signal Sequences
•
•
•
•
N-terminal domain, 15-25 aa’s (longer in G+)
First, region of 3-8 residues with 1-3 +aa
Next, hydrophobic core: 7-15 hydrophobic aa’s
SS cleavage site C-terminal to hydrophobic core
(after Gly/Ala; sometimes more “polar”)
Localization of precursor polypeptides
depends on SS’s and their folding state
• Cytoplasmic proteins stay in cytoplasm
because they lack SS
$They also fold quickly…
• SS target polypeptides to export (or
secretion/Sec) machinery
• Export machinery (Sec apparatus) depends
on unfolded state of polypeptide for
localization
$SS can antagonize folding of protein…
See Fig. 3.12 for characteristics of aa side chains
Bacterial secretion (Sec) apparatus
and post-translational export
Step 1: SecA + SS-precursor;
SecB association
Step 2:
SecA cleaves ATP;
SecEYG pore for
translocation
Essential: SecA, SecEY, Lep
Helpful: SecB, SecG, SecDF
Step 3: repeat ATP-driven SecA cycle
feeding 20-30 aa segments through Sec;
PMF-driven SecEYG cycle;
SS-cleavage by Lep occurs early
Secretion during translation
(Co-translational export)
SRP (signal
recognition
particle) can
associate with
some bacterial
proteins like
cytoplasmic
membrane
proteins during
synthesis. SRP
binds to receptor
near Sec and
facilitates Sec
export into
membrane
Tat Secretion of folded proteins
Secretion during translation
(Co-translational)
SRP is RNA/protein
complex, largely
targets proteins with
very hydrophobic
sequences
(transmembrane
domains) to Sec in
Bacteria. For the
membrane proteins,
no SS cleavage.
Bacterial SRP has
similar structure but
somewhat different
function to euk SRP,
which hasprimary
role targeting SScontaining precursors
to SEC.
Fig. 7.33
Comparing the timing and
apparatuses of export
•
•
•
Bacteria: lots of post and co-translational Sec; distinct SecA
component; Bacteria with other more dedicated posttranslational export machineries such as Tat
Archaea: No SecA but remaining apparatus very similar to
Bacterial Sec; also has Tat export
Eukaryotes: mostly co-translational SEC export; SEC in ER; no
SecA component (Tat apparatus in chloroplasts)
Alternative
apparatus used
in export of
folded bacterial
proteins: TatABC
Tat substrates with
distinct secretion
signals (Twin Arg
Tag); signal
binds TatBC,
then exported via
TatABC, using
PMF