1989 Fiscal Year Final Research Report Summary
Bio-organic Studies on Inhibition of Tubulin Assembly by Mitosis Inhibitors
| Project/Area Number |
63470127
|
|---|
| Research Category |
Grant-in-Aid for General Scientific Research (B)
|
| Allocation Type | Single-year Grants |
|---|
| Research Field |
Chemical pharmacy
|
|---|
| Research Institution | Institute of Applied Microbiology, The University of |
|---|
Principal Investigator |
IWASAKI Shigeo Professor. Institute of Applied Microbiology, 応用微生物研究所, 教授 (00013326)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
FURUKAWA Jun Assistant. Institute of Applied Microbiology, 応用微生物研究所, 助手 (80111574)
|
|---|
| Project Period (FY) |
1988 – 1989
|
| Keywords | tubulin / rhizoxin / maytansine site / 100th amino acid / Photoaffinity label / strucuture-activity |
|---|
| Research Abstract |
This research program was aimed to understand the interaction between tubulin and the mitosis inhibitors, and also to clarify structure and function of tubulin. Previously we have demonstrated with porcine brain tubulin that there exists the rhizoxin/maytansine binding site on tubulin which is different from those for colchicine and for vinblastine.
In this research program, we could show the followings: EBI cross-linking to cystein residues on bovine brain beta-tubulin to form beta^s was strongly inhibited by rhizoxin as maytansine, whereas it was not inhibited by colchicine and was only partially inhibited by vinblastine. This coincides with the previous finding obtained with porcine brain tubulin. In a variety of fungi, the resistance to rhizoxin and ansamitocin P-3 (a maytansinoid) did not cross to the resistance to benzimidazoles. This also agrees with the above results, since benzimidazoles are known to bind to the colchicine site. In Aspergillus nidulans, amino acid sequences of beta-tubulins were determined for wild type (rhizoxin/maytansine sensitive) and rhizoxin (and maytansine) resistant strains. The difference between their sequences lied only in the 100th amino acid; the sensitive tubulin had Asn^<100> and the resistant tubulin had Ile^<100>. Interestingly, Schizosacchromyces pombe and Saccharomyces cervisiae had Ile^<100> and Val^<100> respectively in their beta-tubulins, and they are insensitive to rhizoxin. Replacement of these amino acids to Asn^<100> by site-directed mutagenesis conferred rhizoxin sensitivity.
On the other hand, the side chain structure of rhizoxin was modified to study the structural requirement for binding to tubulin. A variety of derivatives were prepared and among them, a derivative was chosen for photo-affinity labeling to tubulin. Likewise, another derivative for photo-affinity labeling to rhizoxin/maytansin site was prepared also from maytannsinoid.
|
