|Budget Amount *help
¥2,400,000 (Direct Cost : ¥2,400,000)
Fiscal Year 1999 : ¥900,000 (Direct Cost : ¥900,000)
Fiscal Year 1998 : ¥1,500,000 (Direct Cost : ¥1,500,000)
As a step toward determination of the structure of the baseplate of bacteriophage T4 in atomic resolution, three precursor structures were chosen for crystallization, One of them was the initiation complex of the wedge of the baseplate, gp10-gp11 complex. It is considered to constitute the tail pins at the vertices of the hexagonal baseplate, which protrude from the bottom of the baseplate. Based on SDS-PAGE, Edman degradation of the complex and analytical ultracentrifugation, it was shown that the-complex is a hetero-hexamer with the subunit composition of (gp10)ィイD23ィエD2 (gp11)ィイD23ィエD2. Crystal of the complex suitable for X-ray analysis has not been obtained although small crystalls have been found. On the other hand, the second target for X-ray crystal analysis, i.e., gp5-gp27 complex, has been crystallized. Based on the results of sedimentation equilibrium and SDS-PAGE, the subunit structure of the complex was concluded to be hetero-nanomer, (gp5)ィイD23ィエD2(gp27)ィイD26ィエD2. The crystal yielded the resolution of approximately 2.5 A. At present, efforts are made to obatain isomorphous crystal containing selenomethionine. Gp5 is a structurally essential component, yet possesses lysozyme domain which locally digests peptidoglycan of the host E. coli upon infection so that the tail tube can reach the inner membrane. Concerning the third target protein, crystal has not been obtained, but we have recently succeeded in expressing a mutant protein which lacks the N-terminal seven residues that contain five negative charges. The mutant protein was shown to retain the native secondary and oligomeric structure. We plan to examine the possibility of crystallization of the mutant protein.