Se-Met Protein

nakagawa at kekvax.kek.jp nakagawa at kekvax.kek.jp
Wed Nov 10 23:24:10 EST 1993


We have been tring to make a Selenomethyonl protein, but we could get.
We will be very happy if someone has any idea to obtain Se-Met protein.

What we had is:

We expressed, purified and successfully crystallized lymphokine
protein, from  E.coli, BL21(DE3)LysS using T7 phage system.  For X-ray
analysis, we are planning to exchange methinine with selenomethionine.
Unfortunately, methionine auxotroph DL41 is the completely different
system from that we had used. Therefore, we reconstructed a vector
suitable for the E.coli DL41, to which the vector inserted with cDNA
clone of the lymphokine was introduced.  However, DL41, transformed with the
vector, did not express the lymphokine. On the other hand, the E.coli
transformed with a fusion protein with maltase was successfully
expressed.  There may be several possible factors affecting the
unsuccessful expression of human lymphokine in DL41; The vector (pkk223)
we used may not be adequate for the expressionsystem or the lymphokine
itself may be toxic for DL41.  Thus, we are now trying othervectors for
the lymphokine expression, but we still do not reach to the
satisfactoryresults.

Atsushi Nakagawa

-----------------------------
Photon Factory, National Laboratory for High Energy Physics
1-1 Oho Tsukuba, Ibaraki, 305, JAPAN

Tel : 81-298-64-1171
Fax : 81-298-64-2801
e-mail : nakagawa at kekvax.kek.jp




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