Large-scale preparation of biologically active measles virus haemagglutinin expressed by attenuated vaccinia virus vectors.
A procedure described here allows the efficient and rapid purification of histidine-tagged measles virus haemagglutinin that is synthesized under the control of powerful promoters (PSFJ1-10 and PSFJ2-16) of the highly attenuated vaccinia virus (VV) strain LC16mO. A single affinity chromatography step purifies recombinant haemagglutinin proteins from the lysates of cells infected with the recombinant VVs. The recovery and purity are both very high (a yield of 0.5-2.8 mg/10(8) cells and purity of >94-98%), indicating that this procedure is approximately 400 times more efficient than the conventional methods used to prepare haemagglutinin. The haemagglutinins are correctly transported to the cell surface and have haemadsorption activity. Moreover, the recombinant haemagglutinin proteins cooperate with the measles virus fusion protein to elicit cell fusion activity. In addition, the antibody titres against measles virus, as measured by enzyme-linked immunosorbent assay using the purified haemagglutinin as the capture antigen, correlated closely with neutralization test titres (R(2) = 0.84, p < 0.05), indicating the preservation of immunologically relevant antigenicity. Such recombinant haemagglutinin preparations will be useful in diagnostic tests that measure functional anti-measles immunity and investigate the biological functions and structure of the haemagglutinin.
Kidokoro
Minoru
M
Department of Research and Development, Chiba Serum Institute, 2-6-1 Kohnodai, Ichikawa, Chiba 272-0827, Japan. kidcolor@ga2.so-net.ne.jp
Aoki
Atsuko
A
Horiuchi
Kiyoshi
K
Shida
Hisatoshi
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D006386:Hemagglutination Tests; D006389:Hemagglutinins, Viral / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D006801:Humans; D008459:Measles virus / Q000235:genetics*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D014616:Vaccinia virus / Q000235:genetics*; D014760:Viral Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
D006389:Hemagglutinins, Viral; D011993:Recombinant Fusion Proteins; D014760:Viral Fusion Proteins; C032397:hemagglutinin protein G, measles virus