The pentenyl arm fits the hydrophobic grooves between subunits via hydrophobic interaction. It is highly likely that K-10256, K-10487 and K-10552 interact with subunits in a similar manner. Taken together, these results provide the molecular basis of HPDs�� ability to inhibit all three catalytic subunits at the same time. To gain mechanistic insights, we compared the Ancitabine (hydrochloride) supplier binding mode of K-7174 with that of bortezomib using X-ray crystallographic data. As shown in Fig. 4B, three molecules of K-7174 bind to the active pockets of the ?1, ?2 and ?5 subunits along hydrophobic grooves in the direction of the ?7, ?1 and ?4 subunits, respectively. In contrast, bortezomib is attached to the ?5 subunit by a hydrogenbond network composed of Ala49, Thr21 and Gly44. Mutations of amino acids within or near the bortezomibbinding pocket in the ?5 subunit, such as Ala49, Thr21, Met45 and Cys52, were reported to cause bortezomib resistance by reducing the affinity to the drug. Among them, Ala49 makes a direct hydrogen bond to bortezomib, explaining why this position is most frequently mutated in bortezomib-resistant cells. Furthermore, a Cys52Phe or Met45Val substitution results in a steric clash between the side chains of these two residues, leading to repulsion of bortezomib from the binding pocket. In contrast, these mutations should not affect the affinity of K-7174 to ?5 subunit, because the binding site of K- 7174 is spatially distinct from the bortezomib-binding pocket. Because K-7174 appears to inhibit proteasome activity with a distinct mode from bortezomib, it is anticipated that K-7174 is effective for bortezomib-resistant cells. Previous studies revealed that a mutation of the PSMB5 gene at NMS-873 biological activity nucleotide position 322, which corresponds to the substitution of Ala49 to Thr, induced conformational changes in the bortezomib-binding pocket of ?5 subunit and was responsible for acquired bortezomib resistance in T-cell acute lymphoblastic leukemia and myeloid leukemia cells. Recently, Ri et al. reported the establishment of bortezomib-resistant MM cell lines by transduction with G322A-mutated PSMB5 cDNA. Taking the same approach, we established three mutant sublines from RPMI8226 cells lentivirally transduced with mutated PSMB5 along with a marker gene VENUS. As contro
