Intracellular production of a major cytomegalovirus antigenic protein in the methylotrophic yeast Pichia pastoris.
We have previously shown that single or multiple epitopes of the major human cytomegalovirus (HCMV) antigens, produced as fusion proteins in prokaryotes can be valuable diagnostic material in the serology of HCMV infection. In this work we moved to a eukaryotic system, to produce one of the most immunogenic HCMV antigens, ppUL44 (also called pp52 due to its apparent molecular size on acrylamide gels), as a non-fusion protein, in an attempt to eliminate some non-specific reactivity of human sera with bacterial carrier proteins. We expressed the DNA encoding ppUL44 in a highly efficient expression system based on the methylotrophic yeast, Pichia pastoris. Good levels of intracellular, soluble pp52 were produced. We observed an indistinguishable pattern of the yeast pp52 from the viral native protein in immunoblotting and a good reactivity with human sera.
Battista
M C
MC
Istituto di Microbiologia, Policlinico S. Orsola, University of Bologna, Italy.
Bergamini
G
G
Campanini
F
F
Landini
M P
MP
Ripalti
A
A
D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D000956:Antigens, Viral / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D003587:Cytomegalovirus / Q000235:genetics* / Q000276:immunology; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000276:immunology; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D024101:Mitochondrial Proteins; D010088:Oxidoreductases / Q000235:genetics; D010843:Pichia / Q000235:genetics*; D010940:Plant Proteins; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D014170:Transformation, Genetic; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology; D000099188:Alternative Oxidase
D000914:Antibodies, Viral; D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; D010088:Oxidoreductases; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D000099188:Alternative Oxidase; C072217:ICP36 protein, Cytomegalovirus