Phylogenetically more conservative epitopes among monoclonal antibody‐defined antigenic sites of human transferrin are involved in receptor binding

Bártek, Jiří; Viklický, V.; Stratil, A.

doi:10.1111/j.1365-2141.1985.tb07330.x

Cited by 20 publications

(11 citation statements)

References 17 publications

(12 reference statements)

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“…Of great interest is the interaction of antibody HTF.14 with transferrin because this antibody is capable of blocking Tf binding to its receptor. Like Bartek et al (1985) we find that the binding of HTF.14 to Tf of various species roughly parallels the ability of the Tf to be bound by human receptor.…”

Section: Discussionmentioning

confidence: 82%

“…7) shows that bovine Tf is capable of delivering the iron needed for cellular proliferation. Bartek et al (1985) report that bovine Tf, at a concentration of 5 pglml, had no effect on the growth of MOLT-3 cells in Tf-supplemented serumfree medium. Our data confirm that growing cells in medium containing a low concentration of bovine Tf (8 pglml) results in suboptimal growth.…”

Section: Discussionmentioning

confidence: 96%

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Comparative studies of the binding and growth‐supportive ability of mammalian transferrins in human cells

Penhallow

Brown-Mason

Woodworth

1986

Journal Cellular Physiology

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The ability of human-derived cells in culture to bind, remove iron from, and grow in the presence of transferrins (Tf) isolated from the sera of species commonly included in tissue culture medium was investigated. Kinetic studies on HeLa cells reveal apparent first-order association rate constants of 0.43 min-1 for human Tf and 0.15 min-1 for equine Tf. Labeled chicken ovo-Tf and fetal bovine Tf were not recognized by the HeLa cells. Competition experiments with HeLa cells that use either isolated Tf or parent serum confirm these findings. Equilibrium binding experiments performed on HeLa cells at 37 degrees C in the presence of 2,4-dinitrophenol to prevent iron removal indicate 1 X 10(6) Tf bound/cell with a dissociation constant (K'D) of 28 nM for human Tf and 182 nM for equine Tf. Equilibrium binding performed at 0 degrees C to prevent endocytosis reveals 4.1-6.7 X 10(5) Tf binding sites/cell with a K'D of 8.3 nM for human Tf and 41.5 nM for equine Tf. Parallel experiments in normal human diploid fibroblast-like MRC-5 cells indicate expression of 0.82-2.78 X 10(5) Tf binding sites/cell with a K'D of 8.2 nM for human and 39.1 nM for equine Tf. Thus, the results of equilibrium binding studies of a more differentiated cell type are consistent with those found for HeLa cells. Fetal bovine Tf was found to compete weakly with labeled human Tf for human receptor on HeLa cells in a soluble receptor assay, with an approximately 500-fold excess needed to reduce binding to half maximal. Iron uptake experiments show an iron donating hierarchy where human greater than horse greater than calf, suggesting that the rate of iron uptake depends on the affinity of receptor for transferrin. Growth experiments involving HeLa cells in chemically defined serum-free medium demonstrate that bovine Tf will support growth as well as human Tf, but at concentrations much higher than are required of human Tf.

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Section: Discussionmentioning

confidence: 82%

Section: Discussionmentioning

confidence: 96%

Comparative studies of the binding and growth‐supportive ability of mammalian transferrins in human cells

Penhallow

Brown-Mason

Woodworth

1986

Journal Cellular Physiology

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“…(25)(26)(27)(28) This may explain the lack of effect of TRC-2 (which shows no interspecies cross-reactivity) in Tf h -TfR interactions; yet the apparent location of TRC-2 specific epitope in the first of the three conserved regions (i.e., homology block A) (33,35) suggests a precaution in such a correlative approach.…”

Section: Discussionmentioning

confidence: 95%

“…The majority of transferrin-specific MAbs previously reported, (8,(23)(24)(25)(26)(27)(28)(29) has been shown to inhibit Tf h -binding to its receptor and hereby also cell growth. TRC-2 differs from these MAbs in respect to its effect on Tf-action, and its use is hence apt to provide new insight into the mode of action of Tf.…”

Section: Discussionmentioning

confidence: 98%

Partial Characterization of the Human Serum Transferrin Epitope Reactive with the Monoclonal Antibody TRC-2

Öztürk

Çirakoğlu

Bermek

2003

Hybridoma and Hybridomics

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A murine monoclonal antibody (MAb) (TRC-2) specific for human serum transferrin (Tf(h)) was developed. This antibody was depressive on cell growth in serum-free medium in the presence of limiting amounts of Tf(h), but it did not inhibit the binding of Tf(h)-alkaline phosphatase (AP) conjugate to the Tf-receptor (TfR) in a cellular enzyme-linked immunosorbent assay (CELISA) system. On the other hand, the immune complex Tf(h)-TRC-2 was implicated to bind to the receptor in indirect CELISA. Moreover, the detectability of Tf(h)-TfR on the cell surface via Tf-bound TRC-2 suggested that the antibody may inhibit the rapid internalization of this complex. To map the TRC-2-specific epitope, Tf(h) was subjected to proteolytic degradation following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. The treatment with trypsin gave rise to, among others, a fragment of about 42 kDa, which was reactive with TRC-2. Through sequence analysis by automated Edman degradation, the N-terminal sequence of the 42 kDa-tryptic fragment was aligned to the N-terminus of mature transferrin (VPDKTVR). The N-terminal sequence of an immunoreactive CNBr-fragment of about 13 kDa was, in turn, identical with the sequence (NQLRGKK) corresponding to the residues 110-116 on Tf(h).

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“…Moreover, antibodies should be a useful tool to search for such evolutionarily conserved regions. Examples of successful applications of this approach to other systems include detection of a domain on human and guinea pig Ia antigens that may have a regulatory function in T-cell activation (28), and identification of a domain on human transferrin that appears to be involved in receptor binding (1).…”

mentioning

confidence: 99%

Identification of a Highly Conserved Domain on Phytochrome from Angiosperms to Algae

Cordonnier

Greppin²,

Pratt³

1986

Plant Physiol.

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A monoclonal antibody (Pea-25) directed to phytochrome from etiolated peas (Pisum sativam L., cv Alaska) binds to an antigenic domain that has been highly conserved throughout evolution. Antigenic crossreactivity was evaluated by immunoblotting sodium dodecyl sulfate sample buffer extracts prepared from lyophilized tissue samples or freshly harvested algae. Pea-25 immunostained an approximately 120-kilodalton polypeptide from a variety of etiolated and green plant tissues, including both monocotyledons and dicotyledons. Moreover, Pea-25 immunostained a similarly sized polypeptide from the moss Physcomitrella, and from the algae Mougeotia, Mesotaeniwm, and Chlamydomonas. Because

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Phylogenetically more conservative epitopes among monoclonal antibody‐defined antigenic sites of human transferrin are involved in receptor binding

Cited by 20 publications

References 17 publications

Comparative studies of the binding and growth‐supportive ability of mammalian transferrins in human cells

Comparative studies of the binding and growth‐supportive ability of mammalian transferrins in human cells

Partial Characterization of the Human Serum Transferrin Epitope Reactive with the Monoclonal Antibody TRC-2

Identification of a Highly Conserved Domain on Phytochrome from Angiosperms to Algae

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