1.Modification of HPV type 16 E6 and E7 genes, and analysis of stability and immunogenicity of the modified proteins.
Huijun ZHI ; Liqun HAN ; Jiao REN ; Houwen TIAN ; Weifing LUO ; Yu LIANG ; Li RUAN
Chinese Journal of Experimental and Clinical Virology 2002;16(2):124-127
BACKGROUNDTo select the mutants of HPV type 16 E6 and E7 genes suitable for construction of vaccine for treatment of cervical cancer.
METHODSE6 and E7 genes were modified by site-directed mutagenesis. Several recombinant vaccina viruses were constructed by inserting the E6 or E7 mutants into the genome of vaccina virus Tiantan strain and employed to study their antigenicity.
RESULTSWestern blot assay showed that the E6 ?mutant? with substitution of Gly for Leu at amino acid site 50 and E7 mutant with substitution of Gly for Cys-24 and Glu-26 had no effect on their stability and antigenicity, but change of the Cys at position 91 of E7 dramatically reduced its stability and antigencity. Conclusion The results confirmed that the Zinc-finger structure at the E7 C-terminal? plays an important role in the integrity and stability of E7 protein.
Animals ; Antibodies, Viral ; biosynthesis ; Female ; Mice ; Mice, Inbred C57BL ; Mutagenesis, Insertional ; Oncogene Proteins, Viral ; genetics ; immunology ; Papillomaviridae ; genetics ; Papillomavirus E7 Proteins ; Repressor Proteins ; Vaccinia virus ; immunology ; Zinc Fingers
2.Specific bovine antibody response against a new recombinant Cryptosporidium parvum antigen containing 4 zinc-finger motifs.
Dirk C DE GRAAF ; Hans DE CONINCK ; Franz PETRY ; Ilka B EECKHOUT ; Johan E PEETERS
The Korean Journal of Parasitology 2002;40(1):59-64
A Cryptosporidium parvum sporozoite and oocyst lambda gt11 cDNA library was screened with a hyperimmune rabbit serum that was developed against insoluble fragments of ultrasonicated oocysts. A clone named Cp22.4.1 encoding a protein of 231 amino acids with 4 zinc-finger domains characterized by a Cys-X2-Cys-X4-His-X4-Cys motif was isolated and characterized. There was a complete match between the sequencing data of the coding region of Cp22.4.1 and the corresponding gene at chromosomal level. Cloning in a pBAD-TOPO-TA expression vector permitted to evaluate the antigenicity of the recombinant His-tagged antigen. This antigen was recognized by 2 out of 5 sera from Cryptosporidium immune calves and not by sera from parasite naive animals.
Amino Acid Sequence
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Animals
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Antibodies, Protozoan/*blood
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Antigens, Protozoan/chemistry/*immunology
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Base Sequence
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Cattle/*immunology
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Cryptosporidium parvum/*immunology
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Molecular Sequence Data
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Protozoan Proteins/chemistry/genetics
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Rabbits
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Recombinant Proteins
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Zinc Fingers/genetics/*immunology
Result Analysis
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