Expression of adenovirus type 12 Elb 58-kDa protein in Escherichia coli and production of antibodies raised against a 58-kDa::β-galactosidase fusion protein

Schughart, Klaus; Wilcken‐Bergmann, Brigitte von; Esche, Helmut

doi:10.1016/0378-1119(87)90005-9

Cited by 4 publications

(4 citation statements)

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“…The results with the del 620 sera (sera 23 to 25) indicate the likelihood of a major B cell epitope within amino acids 21 to 95 (base pairs 1904 to 2129) of the 54K protein. Taken together it may be that the antisera discussed in this paper and those obtained by Schugart et al (1987) recognize a common epitope between amino acids 64 and 95 of the 54K protein. Further work would be needed to confirm this.…”

mentioning

confidence: 92%

“…Previous work to determine whether polyclonal antisera could be raised to Adl2 54K-fl-galactosidase fusion proteins (Schugart et al, 1987) showed that only very weak antibodies could be raised against the protein containing amino acids 158 to 205 (present in the 54M fusion protein described in this paper). However, strong antibodies were raised against a protein containing amino acids 64 to 205 of the 54K protein, the first 93 amino acids of which overlap with the 54N fusion protein documented in this paper.…”

mentioning

confidence: 99%

“…The smaller E1B protein is 163 amino acids in length and has an Mr of 19K; the larger E1B protein is 482 amino acids in length and has an Mr of 54K. The generation of antibodies for the detection and study of Ad transforming proteins has previously been accomplished by the use of synthetic peptides (Lucher et al, 1984), bacterially expressed proteins (Scott et al, 1984;Schugart et al, 1987), or by the use of tumourbearer serum (TBS) (Rowe et al, 1983). Relatively few studies have been carried out to define the degree or quality of humoral responses to individual E 1 proteins in tumour-bearing animals.…”

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confidence: 99%

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The use of beta-galactosidase fusion proteins encoding the early region 1 transforming proteins of adenovirus type 12 to examine the humoral response in tumour-bearing animals

Merrick

Grand

Brown

et al. 1991

Journal of General Virology

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Sera from 26 rats bearing tumours induced by wildtype (wt) and mutant human adenovirus type 12 (Ad 12), or by cells transformed with these viruses, were analysed for antibodies against the early region 1 (El) transforming proteins. Six Adl2-fl-galactosidase fusion proteins encoding different regions of the Ad 12 E 1 proteins were constructed. The sera from the tumourbearing animals reacted most strongly with the fusion protein encoding the N terminus of the E1A protein.Tumour-bearing rats exposed to the E1B 54K and 19K proteins showed strong reactions with the N terminus of the 54K protein and the C terminus of the 19K protein. Monospecific polyclonal antisera were raised against five of the fusion proteins by immunization of rats and rabbits; these sera cross-reacted with the purified native protein. No antibodies could be obtained which recognized a fusion protein containing amino acids 136 to 268 of the 54K protein. The fusion proteins were also used to purify monospecific antisera from tumour-bearer sera using affinity chromatography.

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confidence: 92%

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confidence: 99%

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confidence: 99%

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The use of beta-galactosidase fusion proteins encoding the early region 1 transforming proteins of adenovirus type 12 to examine the humoral response in tumour-bearing animals

Merrick

Grand

Brown

et al. 1991

Journal of General Virology

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“…The desired protein band was cut from the membrane and subjected to amino acid sequencing in an Applied Biosystems Model 470A gas phase sequencer. Total protein was isolated from E. coli and A. tumefaciens as described by Schughart et al (1987). Total protein was isolated from tobacco calli as described by Powell Abel et al (1986).…”

Section: Methodsmentioning

confidence: 99%

The Nucleotide Sequence of a Soybean Mosaic Virus Coat Protein-coding Region and Its Expression in Escherichia coli, Agrobacterium tumefaciens and Tobacco Callus

Eggenberger

Stark

Beachy

1989

Journal of General Virology

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SUMMARYA DNA complementary to the T-terminal 1168 nucleotides of the genome of the N strain of soybean mosaic virus (SMV) has been cloned and sequenced, cDNA sequence and coat protein analyses indicate that the SMV coat protein-coding region is at the 3' end of the genome, and that the coat protein is processed from a larger protein. The coat protein-coding sequence is predicted to be 795 nucleotides in length, encoding a protein of 265 amino acids with a calculated Mr of 29 857. The 3' untranslated region is 259 nucleotides in length and is followed by a polyadenylate tract. The SMV coat proteincoding region, along with a small amount of upstream sequence, has been expressed in Escherichia coli as a/~-galactosidase fusion protein. The size of the protein was less than predicted for the fusion protein, suggesting processing in E. coli. The coat protein-coding region has also been expressed in Agrobacterium tumefaciens and transgenic tobacco callus as an unfused protein under the control of the cauliflower mosaic virus 35S promoter. The coat protein produced in transgenic tobacco callus had an electrophoretic mobility identical to that of SMV coat protein and constituted approximately 0.05~ (w/w) of the total extracted protein.

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Rabbit antisera to the variable region domains of an anti-α(1 → 6)dextran using E. coli-produced VL and VH fusion proteins as immunogens

Black

Kabat

Morrison

1990

Journal of Immunological Methods

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Expression of adenovirus type 12 Elb 58-kDa protein in Escherichia coli and production of antibodies raised against a 58-kDa::β-galactosidase fusion protein

Cited by 4 publications

References 29 publications

The use of beta-galactosidase fusion proteins encoding the early region 1 transforming proteins of adenovirus type 12 to examine the humoral response in tumour-bearing animals

The use of beta-galactosidase fusion proteins encoding the early region 1 transforming proteins of adenovirus type 12 to examine the humoral response in tumour-bearing animals

The Nucleotide Sequence of a Soybean Mosaic Virus Coat Protein-coding Region and Its Expression in Escherichia coli, Agrobacterium tumefaciens and Tobacco Callus

Rabbit antisera to the variable region domains of an anti-α(1 → 6)dextran using E. coli-produced VL and VH fusion proteins as immunogens

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