Transport Flashcards
Transport of molecules in general
- Simple diffusion (nonpolar components only, down concentration gradient)
- Facilitated diffusion (down electron gradient)
- primary active transport (against electron gradient, needs ATP)
- secondary active transport(against electron gradient, driven by ion moving down its gradient)
- Ion channel (down electrochemical gradient by ligand or ion)
- Ionophoremediated ion transport (down electrochemical gradient)
How are metals transported into the cell ?
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Active transport:
Protein-Mediated active Transport
e.g: Iron, copper are transported into cell through active transport processes gacilitated by specific membrane transporte proteins. -
Passive Diffusion:
Non-specific Channels:
e.g: potassium and sodium can enter cells through non-specific ion channels, these allow ions to move down their concentration gradient -
Endocytosis:
Receptor-Mediated Endocytosis
for large metal-containing molecules or complexes, involves formation of vesicles at the cell membrane. -
Metal Ionophores
are carrier molecules that can bind to metal ions and transport the across cell membranes: Ionophores act as carriers that facilitate the movement of metal ions the cell. -
Metal Chelators:
cells may produce specific molecules = chelators that can bind to metal ions an dtransport the into the cell. This involves the formation of stable complexes between the chelator and the metal ion, enhancing its solubility and transport. e.g: Ferritin, Transferrin and EDTA
- Fe3+ is bound to siderophores and transported into cell (siderophores are small organic molecules produced by microorganisms under iron-limiting conditions which enhance the uptake of iron to the microorganism)
- the oxyanion molybdate (MoO42-)is transported into the cell cia ABC-Transporter (primary active transportes, that involve the direct use of ATP to transport substrates against their concentration gradient)
The ABC Transporter (ATP Binding Cassette)
- largest family of membrane transport proteins.
- They use the energy derived from ATP hydrolysis to transport a variety of small molecules including: Amino acids, sugars, inorganic ions, peptides
- ABC importers require a substrate binding protein that captures the protein and feeds them to external face of the transporter
- ABC transporters also catalyze the flipping of lipids between monolayers in membranes
- consists of 4 protein domains:
1. two hydrophobic membrane-spanning domains (MSDs) that form the channel across the membrane
2. two hydrophilic nucleotide-binding domains (NBDs) that interact at the cytoplasmic surface to supply the energy for active transport
Explain the ABC Transporter function:
Explain ABC-Transporter on the Example: The high affinity molybdate transporter Archeologus fulgidus
- construction:
1. Binding protein (ModA)
2. Trans membran domain TMD (ModB)
3. Nucleotide binding domain NBDs (ModC) - ModA: Molybdate binds to ModA, the periplasmic binding protein.
- ModB: The Transmembran protein has 6 transmembrane helices, these helices form the gate and are highly conserved
- ModC: the nucleotide binding domain
conformational changes observed,
–> binding ATP promotes an outward conformation
–> hydrolysis and dissociation of the products promtes an inward-facing conformations
How are proteins transported to the periplasm?
- Proteins usually tarnsported post-translational
- Membrane proteins are transportert co-translational
- SRP-Pathway = Membrane proteins
- Sec-Pathway = secretory Proteins, unfold
- Tat-Pathway = secretory proteins, folded
Transport of unfolded proteins to the membrane and the periplasm
the Sec-System : secretory system
- protein translocation system
- integral membrane heterotrimer SecYEG (Translocon), translocation channel in the inner membrane of bacteria
- peripheral homodimeric ATPase SecA, provides the energy needed for translocation of proteins across the membrane.
- molecular chaperone SecB helps to keep pre-secretory proteins in a translocation-component state.
- SecDFYajC Complex, comprised of SecDF a membrane protein complex associated with SecYEG translocon, contributed to polypeptidechain movement across membrane
- proteins targeted by the Sec-system are characterized by a leader peptide
How does the Sec-system works?
- Preproteins are bound to SecB in the cytosol
- SecA and SecB build a complex, SecB+Preprotein is targeted to SecYEG translocation site
- SecA binds to ATP and allows this interaction allows the preprotein complex to insert into the membrane
- hydrolysis of ATP releases the preprotein
- in the presence of the proton motive force(PMF), translocation will be completed
*PMF: form of energy generated by the movement of protons (H+Ionen) across a biologial membrane, consists of two components proton gradient and membrane potential * - the unfolded state of the translocated protein is stabilized by molecular chaperones –> SecB
- SecB contains a Channel (Polypeptide binding grooves)
- tetrameric SecB binds the dimeric form of SecA
- SecA functions as an ATP-dependent motor protein and is the central component of the Sec translocase
- ATP is bound and hydrolyze to induce the conformational changes necessary for preprotein translocation
- The channel consists of three integral membrane proteins SecYEG
- SecYEG (the channel) forms a structure with a pore ring of hydrophobic amino acids
- the channel is closed by a plug formed by a short a-helix that folds back into the funnel
(funnel=structure of opening of the channel. SecYEG has a funnel like structure at the entrance of the channel) - signal sequence insertion into the lateral gate widens the central channel, opening and destabilizes the plug, resulting in the opening of a vectorial water-filled-channel
–> c-type cytochromes, hemes are attached to the unfolded apocytochrome in the periplasm, use the Sec-pathway
The SRP (Signale Recognition Particle) Pathway
cellular mechanism involved in the trageting and translocation of proteins across membranes in eukaryotic cells
This pathway ensures that newly synthesized proteins are directed to their appropriate cellular location, such as ER.
- used to for the assembly of cytoplasmic membrane proteins
- SRP binds to Ribosome translating SRP substrates
- interaction with the membrane associated receptor FtsY in an GTP dependent manner
- the translocon occurs via SecYEG complex
- instertion into the membrane via YidC
How do the Sec system and the SRP pathway work together in protein translocation?
*in eukaryotes
- SRP Recognition: SRP recognizes signal sequences on nascent polypeptides during translation.
- SRP Binding and Pause: SRP binds to the signal sequence, leading to a temporary pause in protein synthesis
- SRP Receptor Interaction: SRP-ribosome-nascent chain complex interacts with the SRP receptor on the membrane, often associated with the Sec translocon.
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Sec Translocon-Mediated Translocation:
The SRP-ribosome-nascent chain complex is transferred to the Sec translocon, facilitating translocation of the nascent polypeptide across the membrane.
nascent polypeptide: is a newly syntheisized, partially formed protein chain that is still in the process of being translated from mRNA template.
5.Resumption of Synthesis: Protein synthesis resumes, and the nascent polypeptide is translocated into the ER lumen or another cellular compartment.
The translocation pathway of folded proteins
- Tat Pathway (Twin argenine leader peptides)
- membrane-bound hydrogenase
How are cofactor-containing proteins translocated?
A lot of proteins in the periplasm contain complex metal-cofactors
- more than 40 proteins known that depend on the Tat-Pathway
- proteins that bind FeS clusters, Moco, NiFe cofactors for hydrogenase, multidomain proteins, proteins that depend on fast folding process
- protein transport by the Tat pathway is solely powerd by the electrochemical gradient
Tat-Pathway
TatABC: protein transport system
This pathway is responsible for transporting fully folded or near-fully folded proteins across the cytoplasmic membrane to the periplasmic space or other cellular compartments.
The Tat pathway transports proteins with a specific signal peptide that contains a conserved twin-arginine motif (RR) near the N-terminus of the signal sequence
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TatA Channel Formation:
Feature: TatA forms the channel component of the transporter.
Topology: It has dual membrane topologies, implying dynamic flipping under certain conditions.
Channel Size: The size of the channels depends on the physical dimensions of the substrate being transported. -
TatBC Complex:
Role: Acts as the initial receptor for Tat substrate proteins.
TatB Recognition: TatB recognizes the twin-arginine motif of the signal peptide and has six transmembrane helices. -
Recruitment of TatA:
Process: Once the TatBC complex has recognized a Tat substrate protein, TatA protein can be recruited.
Dependency: Recruitment is dependent on the transmembrane electrochemical gradient. -
TatA Channel Formation (Again):
Function: TatA forms the protein-conducting channel through which the substrate must pass. -
Energetic Cost of Tat Pathway:
Energetically Expensive: The Tat pathway is energetically expensive, requiring approximately 10,000 molecules of ATP per transported protein.
At what stage is the cofactor inserted?
Generally, cofactor insertion occurs after translocation when the protein is in the periplasm, providing an environment conducive to cofactor incorporation and proper folding.
* The exact timing of cofactor insertion can depend on the nature of the substrate protein and the specific cofactors involved.
* Some proteins may already have cofactors embedded in their structures before translocation, while others may acquire cofactors during or after translocation.
