Characterization of cDNA clones for human myeloperoxidase: Predicted amino acid sequence and evidence for multiple mRNA species

Abstract We have isolated 5′ cDNA clones encoding a member of the cellular myosin heavy chain gene family from human leukocytes. The predicted amino acid sequence shows 93% identity to a chicken cellular myosin heavy chain, 76% to chicken smooth muscle, and 40% to human sarcomeric myosin heavy chain. The mRNA is expressed as a 7.4- to 7.9-kb doublet in many nonmuscle cells, and is upregulated in myeloid cell lines on induction from a proliferating to a differentiated state. Antisera raised against a peptide made from the predicted amino acid sequence specifically reacts with a 224-Kd polypeptide in leukocyte cell lines, and the protein is also upregulated during the induction of monocytic and granulocytic differentiation in these cells. The gene for this cellular myosin heavy chain maps to chromosome 22, bands q12.3-q13.1, demonstrating that it is not located in the previously described sarcomeric gene clusters on chromosomes 14 and 17. This cellular myosin heavy chain may be a major contractile protein responsible for movement in myeloid cell lines because no mRNA for sarcomeric myosin heavy chain is detected in these cells.

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Structural features of the low-molecular-mass human salivary mucin

Biochemical Journal ◽

10.1042/bj2870639 ◽

1992 ◽

Vol 287 (2) ◽

pp. 639-643 ◽

Cited By ~ 27

Author(s):

M S Reddy ◽

L A Bobek ◽

G G Haraszthy ◽

A R Biesbrock ◽

M J Levine

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Molecular Mass ◽

Gel Filtration ◽

Structural Features ◽

Cdna Expression Library ◽

Cdna Clones ◽

Expression Library ◽

Mrna Species ◽

Salivary Mucin

The low-molecular-mass human salivary mucin has at least two isoforms, MG2a and MG2b, that differ primarily in their sialic acid and fucose content. In this study, we characterize further these isoforms, particularly their peptide moieties. Trypsin digests of MG2a and MG2b yielded high- and low-molecular-mass glycopeptides following gel filtration on Sephacryl S-300. The larger glycopeptides from MG2a and MG2b had similar amino acid compositions and identical N-terminal sequences, suggesting common structural features between their peptides. An oligonucleotide probe generated from the amino acid sequence of the smaller glycopeptide from MG2a was employed in Northern-blot analysis. This probe specifically hybridized to two mRNA species from human submandibular and sublingual glands. A cDNA clone selected from a human submandibular gland cDNA expression library with antibody generated against deglycosylated MG2a also hybridized to these two mRNA species. In both cases, the larger mRNA was polydisperse, and the hybridization signal was more intense in the sublingual gland. In addition, the N-terminal amino acid sequence of the larger glycopeptide was found to be part of one of the selected MG2 cDNA clones.

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Molecular Characterization of an Avian Astrovirus

Journal of Virology ◽

10.1128/jvi.74.13.6173-6177.2000 ◽

2000 ◽

Vol 74 (13) ◽

pp. 6173-6177 ◽

Cited By ~ 100

Author(s):

Matthew D. Koci ◽

Bruce S. Seal ◽

Stacey Schultz-Cherry

Keyword(s):

Amino Acid ◽

Nucleotide Sequence ◽

Amino Acid Sequence ◽

Animal Species ◽

Genomic Organization ◽

Predicted Amino Acid Sequence ◽

Enteric Disease ◽

Nucleotide Level ◽

Human Astroviruses

ABSTRACT Astroviruses are known to cause enteric disease in several animal species, including turkeys. However, only human astroviruses have been well characterized at the nucleotide level. Herein we report the nucleotide sequence, genomic organization, and predicted amino acid sequence of a turkey astrovirus isolated from poults with an emerging enteric disease.

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Purification and structural characterization of ovine placental lactogen

Journal of Endocrinology ◽

10.1677/joe.0.1260141 ◽

1990 ◽

Vol 126 (1) ◽

pp. 141-149 ◽

Cited By ~ 25

Author(s):

W. C. Warren ◽

R. Liang ◽

G. G. Krivi ◽

N. R. Siegel ◽

R. V. Anthony

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Structural Characterization ◽

Binding Sites ◽

Untranslated Region ◽

Fetal Liver ◽

Radioreceptor Assay ◽

Placental Lactogen ◽

Predicted Amino Acid Sequence ◽

Cdna Clones

ABSTRACT Discrepancies exist in the reported purity and biological activity of ovine placental lactogen (oPL), and little structural characterization has been reported. Ovine PL was purified from fetal cotyledonary tissue (day 100 of gestation) by monitoring activity with a recombinant bovine GH (bGH) liver radioreceptor assay. Two hundred grams of tissue yielded 4·2 mg of oPL, with an ∼ 1000-fold purification of oPL specific activity following initial tissue extraction. The oPL was radioiodinated and used in an ovine fetal liver (day 100 of gestation) radioreceptor assay to examine competitive displacement of oPL, ovine GH (oGH) and ovine prolactin (oPRL). The potency of oPL (ED50 = 0·18 nmol/l; ED50 is the quantity of ligand necessary to displace 50% of specifically bound 125I-labelled oPL) was greater than that of oGH (ED50 = 4·1 nmol/l) and oPRL (ED50 = 1·1 μmol/l) in competing for 125I-labelled oPL-binding sites. Attempts to sequence the NH2 terminus of oPL by vapour-phase sequencing indicated that the NH2 terminus was blocked. Purified oPL was subjected to trypsin and CnBr digestion, and two CnBr and six tryptic peptides were sequenced. The peptide sequences were compared with other PLs, oPRL and bGH for sequence similarity, and found to be most similar to bovine PL (bPL; 68% overall identity) and oPRL (47% overall identity). Complementary DNA (cDNA) clones were isolated for oPL by screening a λZAP cDNA library with a cDNA coding for bPL. Three cDNAs were nucleotide sequenced, and their combined sequence included 41 nucleotides of 5'-untranslated region, the complete coding region of pre-oPL (708 nucleotides) and a portion of the 3' untranslated region (158 nucleotides). The predicted amino acid sequence derived from the nucleotide sequence confirmed homology to bPL (67%) and oPRL (48%). Little amino acid sequence existed with other PLs (≤29%) or GH proteins (≤27%). These results suggest that oPL and oGH are more biologically similar in their ability to compete for fetal liver binding sites, but that oPL is structurally more similar to oPRL. Elucidation of exact structure–function relationships for oPL will, however, require further investigation. Journal of Endocrinology (1990) 126, 141–149

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Cellular myosin heavy chain in human leukocytes: isolation of 5' cDNA clones, characterization of the protein, chromosomal localization, and upregulation during myeloid differentiation

Blood ◽

10.1182/blood.v78.7.1826.bloodjournal7871826 ◽

1991 ◽

Vol 78 (7) ◽

pp. 1826-1833

Author(s):

LE Toothaker ◽

DA Gonzalez ◽

N Tung ◽

RS Lemons ◽

MM Le Beau ◽

...

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Myosin Heavy Chain ◽

Cell Lines ◽

Heavy Chain ◽

Myeloid Cell ◽

Predicted Amino Acid Sequence ◽

Cdna Clones ◽

Granulocytic Differentiation ◽

Human Leukocytes

We have isolated 5′ cDNA clones encoding a member of the cellular myosin heavy chain gene family from human leukocytes. The predicted amino acid sequence shows 93% identity to a chicken cellular myosin heavy chain, 76% to chicken smooth muscle, and 40% to human sarcomeric myosin heavy chain. The mRNA is expressed as a 7.4- to 7.9-kb doublet in many nonmuscle cells, and is upregulated in myeloid cell lines on induction from a proliferating to a differentiated state. Antisera raised against a peptide made from the predicted amino acid sequence specifically reacts with a 224-Kd polypeptide in leukocyte cell lines, and the protein is also upregulated during the induction of monocytic and granulocytic differentiation in these cells. The gene for this cellular myosin heavy chain maps to chromosome 22, bands q12.3-q13.1, demonstrating that it is not located in the previously described sarcomeric gene clusters on chromosomes 14 and 17. This cellular myosin heavy chain may be a major contractile protein responsible for movement in myeloid cell lines because no mRNA for sarcomeric myosin heavy chain is detected in these cells.

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The two beta-tubulin genes of Chlamydomonas reinhardtii code for identical proteins.

Molecular and Cellular Biology ◽

10.1128/mcb.4.12.2686 ◽

1984 ◽

Vol 4 (12) ◽

pp. 2686-2696 ◽

Cited By ~ 115

Author(s):

J Youngblom ◽

J A Schloss ◽

C D Silflow

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Chlamydomonas Reinhardtii ◽

Yeast Cells ◽

Predicted Amino Acid Sequence ◽

Cdna Clones ◽

Beta Tubulin ◽

The Third ◽

Tubulin Genes ◽

Intervening Sequences

The two beta-tubulin genes of the unicellular green alga Chlamydomonas reinhardtii are expressed coordinately after deflagellation and produce two transcripts of 2.1 and 2.0 kilobases. Full-length cDNA clones corresponding to the transcript of each gene were isolated. DNA sequences were obtained from the cDNA clones and from cloned tubulin gene fragments. Both genes contained 1,332 base pairs of coding sequence, with only 19 nucleotide differences between the genes. Because all the differences occurred at the third base position of a codon and did not change the predicted amino acid sequence, we concluded that both beta-tubulin genes code for the same protein of 443 amino acids. The predicted amino acid sequence is 89 and 72% homologous with beta-tubulins from chicken and yeast cells, respectively. Each gene had three intervening sequences, which occurred at identical positions. Although the first two intervening sequences were not conserved between the two genes, the nucleotide sequence of the third intervening sequence was 89% conserved between the genes. The codon usage in the tubulin genes of C. reinhardtii was very biased: only 37 different codons were used. Striking differences occurred between the codons used in these nuclear genes and C. reinhardtii chloroplast genes.

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cDNA cloning of transcription factor E4TF1 subunits with Ets and notch motifs

Molecular and Cellular Biology ◽

10.1128/mcb.13.3.1385-1391.1993 ◽

1993 ◽

Vol 13 (3) ◽

pp. 1385-1391

Author(s):

H Watanabe ◽

J Sawada ◽

K Yano ◽

K Yamaguchi ◽

M Goto ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Amino Acid ◽

Dna Binding ◽

Amino Acid Sequence ◽

Binding Activity ◽

Amino Acid Sequences ◽

Predicted Amino Acid Sequence ◽

Cdna Clones ◽

Dna Binding Activity

E4TF1 was originally identified as one of the transcription factors responsible for adenovirus E4 gene transcription. It is composed of two subunits, a DNA binding protein with a molecular mass of 60 kDa and a 53-kDa transcription-activating protein. Heterodimerization of these two subunits is essential for the protein to function as a transcription factor. In this study, we identified a new E4TF1 subunit, designated E4TF1-47, which has no DNA binding activity but can associate with E4TF1-60. We then cloned the cDNAs for each of the E4TF1 subunits. E4TF1 was purified, and the partial amino acid sequence of each subunit was determined. The predicted amino acid sequence of each cDNA clone revealed that E4TF1-60 had an ETS domain, which is a DNA binding domain common to ets-related transcription factors. E4TF1-53 had four tandemly repeated notch-ankyrin motifs. The putative cDNA of E4TF1-47 coded almost the same amino acid sequences as E4TF1-53. Three hundred and thirty-two amino acids of the N termini of E4TF1-47 and -53 were identical except for one amino acid insertion in E4TF1-53, and they differ from each other at the C terminus. These three recombinant cDNA clones were expressed in Escherichia coli, and the proteins behaved in the same manner as purified proteins in a gel retardation assay. Nucleotide and predicted amino acid sequences were highly homologous to GABP-alpha and -beta, which is further supported by the observation that GABP-specific antibody can recognize human E4TF1.

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cDNA cloning of transcription factor E4TF1 subunits with Ets and notch motifs.

Molecular and Cellular Biology ◽

10.1128/mcb.13.3.1385 ◽

1993 ◽

Vol 13 (3) ◽

pp. 1385-1391 ◽

Cited By ~ 66

Author(s):

H Watanabe ◽

J Sawada ◽

K Yano ◽

K Yamaguchi ◽

M Goto ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Amino Acid ◽

Dna Binding ◽

Amino Acid Sequence ◽

Binding Activity ◽

Amino Acid Sequences ◽

Predicted Amino Acid Sequence ◽

Cdna Clones ◽

Dna Binding Activity

E4TF1 was originally identified as one of the transcription factors responsible for adenovirus E4 gene transcription. It is composed of two subunits, a DNA binding protein with a molecular mass of 60 kDa and a 53-kDa transcription-activating protein. Heterodimerization of these two subunits is essential for the protein to function as a transcription factor. In this study, we identified a new E4TF1 subunit, designated E4TF1-47, which has no DNA binding activity but can associate with E4TF1-60. We then cloned the cDNAs for each of the E4TF1 subunits. E4TF1 was purified, and the partial amino acid sequence of each subunit was determined. The predicted amino acid sequence of each cDNA clone revealed that E4TF1-60 had an ETS domain, which is a DNA binding domain common to ets-related transcription factors. E4TF1-53 had four tandemly repeated notch-ankyrin motifs. The putative cDNA of E4TF1-47 coded almost the same amino acid sequences as E4TF1-53. Three hundred and thirty-two amino acids of the N termini of E4TF1-47 and -53 were identical except for one amino acid insertion in E4TF1-53, and they differ from each other at the C terminus. These three recombinant cDNA clones were expressed in Escherichia coli, and the proteins behaved in the same manner as purified proteins in a gel retardation assay. Nucleotide and predicted amino acid sequences were highly homologous to GABP-alpha and -beta, which is further supported by the observation that GABP-specific antibody can recognize human E4TF1.

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The two beta-tubulin genes of Chlamydomonas reinhardtii code for identical proteins

Molecular and Cellular Biology ◽

10.1128/mcb.4.12.2686-2696.1984 ◽

1984 ◽

Vol 4 (12) ◽

pp. 2686-2696

Author(s):

J Youngblom ◽

J A Schloss ◽

C D Silflow

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Chlamydomonas Reinhardtii ◽

Yeast Cells ◽

Predicted Amino Acid Sequence ◽

Cdna Clones ◽

Beta Tubulin ◽

The Third ◽

Tubulin Genes ◽

Intervening Sequences

The two beta-tubulin genes of the unicellular green alga Chlamydomonas reinhardtii are expressed coordinately after deflagellation and produce two transcripts of 2.1 and 2.0 kilobases. Full-length cDNA clones corresponding to the transcript of each gene were isolated. DNA sequences were obtained from the cDNA clones and from cloned tubulin gene fragments. Both genes contained 1,332 base pairs of coding sequence, with only 19 nucleotide differences between the genes. Because all the differences occurred at the third base position of a codon and did not change the predicted amino acid sequence, we concluded that both beta-tubulin genes code for the same protein of 443 amino acids. The predicted amino acid sequence is 89 and 72% homologous with beta-tubulins from chicken and yeast cells, respectively. Each gene had three intervening sequences, which occurred at identical positions. Although the first two intervening sequences were not conserved between the two genes, the nucleotide sequence of the third intervening sequence was 89% conserved between the genes. The codon usage in the tubulin genes of C. reinhardtii was very biased: only 37 different codons were used. Striking differences occurred between the codons used in these nuclear genes and C. reinhardtii chloroplast genes.

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