Abstract
Ehrlichia chaffeensis, the causative agent of human monocytotropic ehrlichiosis, secretes several effector proteins that bind host DNA to modulate host gene expression. The tandem repeat protein 120 (TRP120), one of the largest effector proteins, has four nearly identical tandem repeat (TR) regions that each consists of 80 amino acids. In addition to playing a role in ehrlichial binding and internalization, TRP120 translocates to the host nucleus where it is thought to function as a transcription factor that modulates gene expression. However, sequence analysis of TRP120 does not identify the presence of DNA-binding or trans-activation domains typical of classical eukaryotic transcription factors. Thus, the mechanism by which TRP120 binds DNA and modulates gene expression remains elusive. Herein, we expressed the TR regions of the TRP120 protein, and characterized its solution structure and ability to bind DNA. TRP120, expressed as either a one or two TR repeat, is a monomer in solution, and is mostly disordered as determined by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. Using NMR spectroscopy, we further show that the 1 TR construct selectively binds GC-rich DNA. Although low pH was required for TRP120 TR-DNA interaction, acidic pH alone does not induce any significant structural changes in the TR region. This suggests that TRP120 folds into an ordered structure upon forming a protein-DNA complex, and thus folding of TRP120 TR is coupled with DNA binding.
Highlights
Ehrlichia chaffeensis, the etiologic agent of human monocytotropic ehrlichiosis (HME), is an obligate intracellular bacterium and a member of the Anaplasmataceae family [1, 2]
To determine if tandem repeat (TR) regions form a dimer that is stable during SDS-PAGE, the oligomeric states of the tandem repeat protein 120 (TRP120) constructs were examined by sedimentation velocity analytical ultracentrifugation
E. chaffeensis tandem repeat protein (TRP) effectors are involved in the activation of host cell pathways, manipulation of gene transcription, and exploitation of post-translational modifications to reprogram the host cell
Summary
The etiologic agent of human monocytotropic ehrlichiosis (HME), is an obligate intracellular bacterium and a member of the Anaplasmataceae family [1, 2]. Transmitted by the lone star tick (Amblyomma americanum), E. chaffeensis infects human mononuclear phagocytes where it replicates forming microcolonies in cytoplasmic membrane-bound vacuoles called. HME presents symptoms ranging from mild febrile illness, malaise, and headache, to life-threatening septic shock, respiratory distress, and renal or hepatic failure in more serious cases, with over half requiring hospitalization and 3% proving fatal [4, 5]. Because the symptoms of E. chaffeensis infection are similar to other undifferentiated febrile illnesses and tick-borne infections endemic to the same geographic areas, HME is often misdiagnosed and underreported [8]
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