Malaria PKG Structure and Activation

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Questions and Answers

What role do autoinhibition mechanisms play in protein kinases?

They increase allosteric interactions.

They prevent unnecessary activation of the enzyme. (correct)

They promote substrate binding.

They enhance phosphorylation efficiency.

Which feature is characteristic of cyclic GMP-dependent protein kinases (cGK) in protozoa?

They may exhibit positive cooperativity. (correct)

They have multiple phosphorylation sites.

They are primarily found in mammalian cells.

They utilize ATP as a primary substrate.

Where is serine 63 located in relation to autophosphorylation in protein kinases?

It is adjacent to the myristoylation site.

It is part of the allosteric binding site.

It is outside the autophosphorylation sequence. (correct)

It is within the catalytic domain.

What is a key characteristic of the PKA RIIβ tetrameric holoenzyme structure?

It reveals insights into allosteric regulation. Signup and view all the answers

In what context is the term 'myristoylation' relevant to protein kinases?

It involves the attachment of a fatty acid for membrane localization. Signup and view all the answers

Which of the following statements is true regarding phosphoproteomics in malaria research?

It focuses on protein phosphorylation patterns in parasites. Signup and view all the answers

What impact does the structure of protein kinases have on their function?

The overall structure dictates substrate specificity. Signup and view all the answers

What is the relationship between cyclic adenosine monophosphate (cAMP) and protein kinase activation?

cAMP acts as a secondary messenger for activation. Signup and view all the answers

What structural feature distinguishes PfPKG-b from PfPKG-a in a monomer form?

Extension of the AIS in the KD Signup and view all the answers

In the context of AGC kinases, what characteristic is noted about the C terminus in Pf PKG structures?

It is a hallmark for AGC kinases Signup and view all the answers

How does the activation loop of Pf PKG differ from that of PKA?

It is in an open conformation and unphosphorylated Signup and view all the answers

Which residue in Pf PKG is indicated to become phosphorylated in the presence of cGMP, Mg2+, and ATP?

T695 Signup and view all the answers

What role does the proline in the PxxP motif play in Pf PKG as compared to PKA?

Facilitates interlobe movement Signup and view all the answers

What role does the N-terminal helix, αAK, play in the structure of PKG?

It interacts with both lobes of the KD and plays a part in maintaining the auto-inhibited state. Signup and view all the answers

In the structures studied, how does the KD of the Pf PKG compare to other kinase structures?

It can be described as partially open Signup and view all the answers

What does the overpass in PfPKG-b indicate about the structural differences?

It may show how PfPKG-a differs structurally as a monomer Signup and view all the answers

Which component is involved in the interaction with helix αAK to maintain the inhibition of PKG?

Salt bridges with specific residue pairs. Signup and view all the answers

What is a significant feature of the kinase domain discovered in all Plasmodium PKG structures?

Presence of a conserved ATP-binding site Signup and view all the answers

What is a primary structural difference between PKA and PKG?

The positioning of helix αAK. Signup and view all the answers

Which specific interactions are crucial for maintaining the open conformation of the KD in PKG?

Salt bridges and π-bonds involving specific amino acids. Signup and view all the answers

In what way do CNB-D cGMP-binding domains interact with αAK?

They create water-mediated contacts for interaction. Signup and view all the answers

What is the significance of the R194C–D241R–D267R triad in relation to PKG?

It replaces a similar triad found in PKA to maintain structural integrity. Signup and view all the answers

What type of interactions does the C tail participate in concerning PKG's structure?

Interactions that promote the auto-inhibited state of the protein. Signup and view all the answers

Which effect does the structural deviation of αAK have on PKG?

It contributes to the protein's auto-inhibitory state. Signup and view all the answers

Study Notes

Malaria Parasite cGMP-Dependent Protein Kinase (PKG) Structure and Activation

Malaria is a global health concern, with ~500,000 annual deaths.
The cyclic guanosine-3',5'-monophosphate (cGMP)-dependent protein kinase (PKG) is a potential target for new antimalarial drugs.
PKG plays a critical role in various stages of the malaria parasite life cycle.
PfPKG and PvPKG structures determined, revealing a previously unknown activation mechanism.
The enzyme is predominantly a monomer, not a dimer as previously thought.

Unique Structural Relay Mechanism

PKG's structure reveals four predicted cyclic nucleotide-binding domains (CNBs) and a kinase domain (KD) arranged in a pentagonal configuration.
An N-terminal autoinhibitory segment (AIS) is crucial for keeping the kinase inactive.
CNBs and KD interact to keep the enzyme inactive
Cooperative binding of cGMP, in a structural relay mechanism, triggers conformational changes around the molecule.
This enables PKG to orchestrate rapid developmental switches in response to cGMP levels.

Significance of Discovery

The previously unknown structure of cGMP-free PKG from malaria parasites provides insight into the activation process.
These structures, coupled to biochemical data, suggest a structural relay activation mechanism involving interdomain communication.
PKG is a promising drug target for malaria.

cGMP-Free PKG Structures

Two orthologs (Plasmodium falciparum and Plasmodium vivax) structures (PfPKG and PvPKG) were elucidated, at ~2.4Å, in an orthorhombic and monoclinic space group, respectively.

Structure-Activity Relationships and Modeling

The N-terminal segment exhibits autoinhibitory characteristics. Removal of the segment showed no negative effect on cGMP-mediated activity
The activation loop is open, an uncommon characteristic for non-phosphorylated AGC kinases
The c terminus exhibits distinctive features, including a critical proline-rich segment conserved among PfPKG orthologs.
The study also confirmed that multiple CNBs bind cGMP, with pCNB-C identified as lacking cGMP-binding capability, unlike the other CNBs.
The crystal structure shows interdomain communication and reveals critical interactions essential for PKG's activation loop.

Molecular Architecture of the Kinase Domain

The kinase domain (KD) exhibits a characteristic bilobal architecture as observed in PKA and other AGC kinases.
The activation loop is present in an open conformation, despite lack of phosphorylation, in a manner not previously observed.
Key structural elements conserved in PKA, like aC-helix, are present, but unique interactions between c-terminal and n-terminal areas of PKG were characterized as contributing to the overall autoinhibited state.

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Description

Explore the structure and activation mechanisms of cGMP-dependent protein kinase (PKG) in malaria parasites. This quiz covers the critical role of PKG in the life cycle of malaria and the discovery of its unique structural relay mechanism. Understand how this knowledge can inform new antimalarial drug development.