Summary Kinesin-2 motors, which are involved in intraflagellar transport and cargo transport along cytoplasmic microtubules, differ from motors in the canonical Kinesin-1 family in having a heterodimeric rather than homodimeric structure and in possessing a three amino acid insertion in their neck linker domain. To determine how these structural features alter the chemomechanical coupling in Kinesin-2, we used single-molecule bead experiments to measure the processivity and velocity of mouse Kinesin-2 heterodimer (KIF3A/B) and the engineered homodimers KIF3A/A and KIF3B/B, and compared their behavior to Drosophila Kinesin-1 heavy chain (KHC). Single motor run lengths of Kinesin-2 were four-fold shorter than Kinesin-1. Extending the Kinesin-1 neck linker by three amino acids led to a similar reduction in processivity. Furthermore, Kinesin-2 processivity varied inversely with ATP concentration. Stochastic simulations of the Kinesin-1 and Kinesin-2 hydrolysis cycles suggest that “front head gating”, in which rearward tension prevents ATP binding to the front head when both heads are bound to the microtubule, is diminished in Kinesin-2. Because the mechanical tension that underlies front head gating must be transmitted through the neck linker domains, we propose that the diminished coordination in Kinesin-2 is a result of its longer and hence more compliant neck linker element.

KIF3A/B, a kinesin involved in intraflagellar transport and Golgi trafficking, is distinctive because it contains two nonidentical motor domains. Our hypothesis is that the two heads have distinct functional properties, which are tuned to maximize the performance of the wild-type heterodimer. To test this, we investigated the motility of wild-type KIF3A/B heterodimer and chimaeric KIF3A/A and KIF3B/B homodimers made by splicing the head of one subunit to the rod and tail of the other. The first result is that KIF3A/B is processive, consistent with its transport function in cells. Secondly, the KIF3B/B homodimer moves at twice the speed of the wild-type motor but has reduced processivity, suggesting a trade-off between speed and processivity. Third, the KIF3A/A homodimer moves fivefold slower than wild-type, demonstrating distinct functional differences between the two heads. The heterodimer speed cannot be accounted for by a sequential head model in which the two heads alternate along the microtubule with identical speeds as in the homodimers. Instead, the data are consistent with a coordinated head model in which detachment of the slow KIF3A head from the microtubule is accelerated roughly threefold by the KIF3B head.

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The accumulation of advanced glycation end products (AGEs) has been reported to be a major contributor to chronic systemic inflammation. AGEs are not efficiently removed by hemodialysis or the kidney of a chronic kidney disease (CKD) patient. The goal of this study was to develop a receptor for AGEs (RAGE)-based bioadsorbent device that was capable of removing endogenous AGEs from human blood. The extracellular domain of RAGE was immobilized onto agarose beads to generate the bioadsorbent. The efficacy of AGE removal from saline, serum, and whole blood; biological effects of AGE reduction; and hemocompatibility and stability of the bioadsorbent were investigated. The bioadsorbent bound AGE-modified bovine serum albumin (AGE-BSA) with a binding capacity of 0.73 ± 0.07 mg AGE-BSA/ml bioadsorbent. The bioadsorbent significantly reduced the concentration of total AGEs in serum isolated from end stage kidney disease (ESKD) patients by 57%. AGE removal resulted in a significant reduction of vascular cell adhesion molecule-1 (VCAM-1) expression in human endothelial cells and abolishment of osteoclast formation in osteoclast progenitor cells. A hollow fiber device loaded with bioadsorbent reduced endogenous AGEs from recirculated blood to 36% of baseline levels with no significant changes in total protein and albumin concentration. The bioadsorbent maintained AGE-specific binding capacity after freeze-drying and storage for 1 year. This approach provides the foundation for further development of sRAGE-based extracorporeal therapies to selectively deplete serum AGEs from human blood and decrease inflammation in patients with diabetes and/or CKD.