Domain structure and three-dimensional model of a group II intron-encoded reverse transcriptase

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Group II intron-encoded proteins (IEPs) exhibit dual reverse transcriptase (RT) and maturase functions essential for intron mobility and RNA splicing.
• The LtrA protein from Lactococcus lactis Ll.LtrB intron possesses an N-terminal RT domain, a maturase domain (domain X), and DNA-binding/endonuclease domains.

Purpose of the Study:

• To investigate the structural and functional characteristics of the LtrA protein, particularly its RT and maturase domains.
• To elucidate the structural basis for LtrA's RNA-binding capabilities and its role in group II intron splicing and reverse transcription.

Main Methods:

• Partial proteolysis using trypsin and Arg-C to identify LtrA cleavage sites.
• Sequence alignments, secondary-structure predictions, and hydrophobicity profiling.
• Three-dimensional structural modeling of LtrA by threading onto HIV-1 RT X-ray crystal structures.

Main Results:

• Proteolysis revealed major cleavage sites within the RT1 region and between the RT and domain X.
• Domain X shows structural similarity to the thumb domain of retroviral RTs.
• Structural models suggest an RNA-binding track analogous to HIV-1 RT and identify conserved regions in splicing-competent LtrA variants involved in RNA binding.

Conclusions:

• The structural model of LtrA provides insights into its proteolytic cleavage sites and template-primer binding.
• Conserved regions in LtrA likely form an extended RNA-binding surface essential for group II intron splicing and reverse transcription.