Synthesis of Heparan Sulfate Hexadecasaccharides and Their Molecular Interaction with Mycobacterial Heparin-Binding Hemagglutinin for the Detection of Mycobacterium tuberculosis.

Heparin-binding hemagglutinin (HBHA), located on the surface of Mycobacterium tuberculosis, binds to heparan sulfate (HS) on respiratory epithelial cells, initiating extrapulmonary dissemination and contributing to the development of latent tuberculosis. Previous characterization suggested that the lysine-rich domain of HBHA may accommodate an HS chain that is approximately twice the length of an octasaccharide. Herein, we prepared eight HS-based hexadecasaccharides through convergent assembly of a precursor, followed by divergent functional group modifications to generate varying sulfonation patterns. The hexadecasaccharide with repeating trisulfonated disaccharide units exhibited the highest binding affinity. The regions of HBHA affected by the interaction were identified by circular dichroism and multidimensional nuclear magnetic resonance analysis. Biotin functionalization of the hexadecasaccharide facilitated attachment to streptavidin-coated magnetic beads. These beads with bound HS displayed the ability to capture mycobacteria via HBHA on an integrated microfluidic chip. Aided by propidium monoazide as a DNA-binding agent and on-chip polymerase chain reaction, a means for diagnosis of tuberculosis is demonstrated.