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The majority of the L1ORF1 structure was solved by X-ray crystallography^[2]. The crystallized part of the protein is a 338-residue, 40 kDa chain composed of : long N-terminal alpha-helical domain, central RNA-recognition and binding domain (RRM) and a C-terminal domain (CTD). The N-terminal residues were not crystallized, but based on previous atomic force microscopy experiments^[3] they are expected to extend the alpha-helix by approximately 50 residues. The domains are connected with two linker regions responsible for structure flexibility. The overall structure of the protein forms an L-shaped pocket formed by N terminal helix and central region with the flexible C-terminal domain “capping” the binding pocket (link to monomer).

Stabilization of the Trimeric Structure

L1ORF1p forms with unusually long alpha helices (residues 111-153), which create a coiled-coil structure. The very long N-terminal helices are stabilized by three structural features:

• Coordination of on the hydrophobic interface inside the coiled-coil structure by three asparagines (Asn142) and three arginines (Arg135). These promote the trimeric state of the coiled-coil domain;
• Externally stabilizing between coiled-coil helices along the hydrophilic face of the helices;
• Water exclusion interactions within the internal hydrophobic face of the helices in the coiled-coil domain.

The trimerization is additionally stabilized on the C-terminal side of the molecule by 1 hydrogen bond between domain, while the CTD regions remain more flexible and do not interact with one another. This might play a significant role in the accommodation of RNA molecules or during retrotransciption.

Nucleic Acid Binding Surfaces

L1ORF1p is an RNA-interacting molecule, although its structure in the RNA-bound state was not crystallized. Careful mapping of protein surface electrostatic potential indicated the likely sites of RNA accommodation. The protein contains two major grooves with positively-charged surface whose affinity to negatively-charged RNA is highest:

• Horizontal cleft between the of each monomer, where the local electrostatic potential reaches up to +15 kT/e;
• Vertical, wide groove between monomers, with somewhat lower positive local electrostatics.

It has been demonstrated by mutagenesis that arginine residues 220, 235 and 261^[4] that are on the surface between the RRM and CTD domains are indispensable for the RNA affinity of the trimer. Therefore, in the RNA-bound state, the RNA is likely wound around the L1ORF1p trunk with the strand passing through the negatively charged vertical and horizontal grooves. So far, however, it is impossible to experimentally verify the structure of the RNA-protein interactions.

Structural Flexibility

The CTD domains are "hinged" to the RRM and coiled-coil domains, and are free to move relative to the rest of the structure. This might be important for reverse transcription, allowing the trimer to unwind the RNA slowly so as to prevent the formation of secondary structure in the transcript. There was found a and an one.

3D Structures of L1 ORF1 Protein

Mus musculus (mouse)

Nuclear Magnetic Resonance: 2JRB, 2LDY, 2W7A

Homo sapiens (human)

X-Ray Crystallography: 2YKO, 2YKP, 2YKQ

See Also

Movie of conformational change

References