The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells

M. A. Kane, P. C. Elwood, R. M. Portillo, Asok Antony, J. F. Kolhouse

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Human KB cells produce two immunologically cross-reactive folate-binding proteins: a particulate cell-associated protein which is solubilized by Triton X-100, and a soluble protein which is released into their growth medium. This compartmentation of these two folate-binding proteins provides a convenient system for studies of their biochemical relationship. The two folate-binding proteins behave similarly to the purified particulate and soluble folate-binding proteins of human milk in analysis by radioactive folate binding, Sephacryl S-200 gel filtration profiles, polyacrylamide gel electrophoresis in either Triton X-100 or sodium dodecyl sulfate, and in Triton X-100 binding based on sucrose density gradient ultracentrifugation in H2O and D2O. The two folate-binding proteins were endogenously labeled by pulsing methionine-starved KB cells with [35S]methionine, and each protein was purified to apparent homogeneity by affinity chromatography at different times during the chase with nonradioactive methionine. The time course of the changes in specific activity (moles of [35S]methionine per mole of folate-binding protein) revealed a more rapid initial rate of synthesis and an earlier maximum in specific activity for the cell-associated folate-binding protein than for the soluble folate-binding protein released into the growth medium. Differences in the levels and specific activities of the two folate-binding proteins of cells exposed to cycloheximide compared with simultaneous controls after pulsing with [35S]methionine suggest that, whereas the cell-associated folate-binding protein is probably produced by de novo protein synthesis, the soluble folate-binding protein seems to be produced from a cellular pool of an already synthesized protein. These results combined with the immunologic cross-reactivity of the two folate-binding proteins strongly suggest a precursor-product relationship between them.

Original languageEnglish (US)
Pages (from-to)15625-15631
Number of pages7
JournalJournal of Biological Chemistry
Volume261
Issue number33
StatePublished - 1986
Externally publishedYes

Fingerprint

KB Cells
Folic Acid
Carrier Proteins
Membranes
Methionine
Octoxynol
Proteins
Affinity chromatography
Ultracentrifugation
Human Milk
Cycloheximide
Growth
Electrophoresis
Affinity Chromatography
Sodium Dodecyl Sulfate
Gel Chromatography
Sucrose
Polyacrylamide Gel Electrophoresis

ASJC Scopus subject areas

  • Biochemistry

Cite this

Kane, M. A., Elwood, P. C., Portillo, R. M., Antony, A., & Kolhouse, J. F. (1986). The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells. Journal of Biological Chemistry, 261(33), 15625-15631.

The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells. / Kane, M. A.; Elwood, P. C.; Portillo, R. M.; Antony, Asok; Kolhouse, J. F.

In: Journal of Biological Chemistry, Vol. 261, No. 33, 1986, p. 15625-15631.

Research output: Contribution to journalArticle

Kane, MA, Elwood, PC, Portillo, RM, Antony, A & Kolhouse, JF 1986, 'The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells', Journal of Biological Chemistry, vol. 261, no. 33, pp. 15625-15631.
Kane, M. A. ; Elwood, P. C. ; Portillo, R. M. ; Antony, Asok ; Kolhouse, J. F. / The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells. In: Journal of Biological Chemistry. 1986 ; Vol. 261, No. 33. pp. 15625-15631.
@article{d94de8b8abdc49acb8ea512530c031b2,
title = "The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells",
abstract = "Human KB cells produce two immunologically cross-reactive folate-binding proteins: a particulate cell-associated protein which is solubilized by Triton X-100, and a soluble protein which is released into their growth medium. This compartmentation of these two folate-binding proteins provides a convenient system for studies of their biochemical relationship. The two folate-binding proteins behave similarly to the purified particulate and soluble folate-binding proteins of human milk in analysis by radioactive folate binding, Sephacryl S-200 gel filtration profiles, polyacrylamide gel electrophoresis in either Triton X-100 or sodium dodecyl sulfate, and in Triton X-100 binding based on sucrose density gradient ultracentrifugation in H2O and D2O. The two folate-binding proteins were endogenously labeled by pulsing methionine-starved KB cells with [35S]methionine, and each protein was purified to apparent homogeneity by affinity chromatography at different times during the chase with nonradioactive methionine. The time course of the changes in specific activity (moles of [35S]methionine per mole of folate-binding protein) revealed a more rapid initial rate of synthesis and an earlier maximum in specific activity for the cell-associated folate-binding protein than for the soluble folate-binding protein released into the growth medium. Differences in the levels and specific activities of the two folate-binding proteins of cells exposed to cycloheximide compared with simultaneous controls after pulsing with [35S]methionine suggest that, whereas the cell-associated folate-binding protein is probably produced by de novo protein synthesis, the soluble folate-binding protein seems to be produced from a cellular pool of an already synthesized protein. These results combined with the immunologic cross-reactivity of the two folate-binding proteins strongly suggest a precursor-product relationship between them.",
author = "Kane, {M. A.} and Elwood, {P. C.} and Portillo, {R. M.} and Asok Antony and Kolhouse, {J. F.}",
year = "1986",
language = "English (US)",
volume = "261",
pages = "15625--15631",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "33",

}

TY - JOUR

T1 - The interrelationship of the soluble and membrane-associated folate-binding proteins in human KB cells

AU - Kane, M. A.

AU - Elwood, P. C.

AU - Portillo, R. M.

AU - Antony, Asok

AU - Kolhouse, J. F.

PY - 1986

Y1 - 1986

N2 - Human KB cells produce two immunologically cross-reactive folate-binding proteins: a particulate cell-associated protein which is solubilized by Triton X-100, and a soluble protein which is released into their growth medium. This compartmentation of these two folate-binding proteins provides a convenient system for studies of their biochemical relationship. The two folate-binding proteins behave similarly to the purified particulate and soluble folate-binding proteins of human milk in analysis by radioactive folate binding, Sephacryl S-200 gel filtration profiles, polyacrylamide gel electrophoresis in either Triton X-100 or sodium dodecyl sulfate, and in Triton X-100 binding based on sucrose density gradient ultracentrifugation in H2O and D2O. The two folate-binding proteins were endogenously labeled by pulsing methionine-starved KB cells with [35S]methionine, and each protein was purified to apparent homogeneity by affinity chromatography at different times during the chase with nonradioactive methionine. The time course of the changes in specific activity (moles of [35S]methionine per mole of folate-binding protein) revealed a more rapid initial rate of synthesis and an earlier maximum in specific activity for the cell-associated folate-binding protein than for the soluble folate-binding protein released into the growth medium. Differences in the levels and specific activities of the two folate-binding proteins of cells exposed to cycloheximide compared with simultaneous controls after pulsing with [35S]methionine suggest that, whereas the cell-associated folate-binding protein is probably produced by de novo protein synthesis, the soluble folate-binding protein seems to be produced from a cellular pool of an already synthesized protein. These results combined with the immunologic cross-reactivity of the two folate-binding proteins strongly suggest a precursor-product relationship between them.

AB - Human KB cells produce two immunologically cross-reactive folate-binding proteins: a particulate cell-associated protein which is solubilized by Triton X-100, and a soluble protein which is released into their growth medium. This compartmentation of these two folate-binding proteins provides a convenient system for studies of their biochemical relationship. The two folate-binding proteins behave similarly to the purified particulate and soluble folate-binding proteins of human milk in analysis by radioactive folate binding, Sephacryl S-200 gel filtration profiles, polyacrylamide gel electrophoresis in either Triton X-100 or sodium dodecyl sulfate, and in Triton X-100 binding based on sucrose density gradient ultracentrifugation in H2O and D2O. The two folate-binding proteins were endogenously labeled by pulsing methionine-starved KB cells with [35S]methionine, and each protein was purified to apparent homogeneity by affinity chromatography at different times during the chase with nonradioactive methionine. The time course of the changes in specific activity (moles of [35S]methionine per mole of folate-binding protein) revealed a more rapid initial rate of synthesis and an earlier maximum in specific activity for the cell-associated folate-binding protein than for the soluble folate-binding protein released into the growth medium. Differences in the levels and specific activities of the two folate-binding proteins of cells exposed to cycloheximide compared with simultaneous controls after pulsing with [35S]methionine suggest that, whereas the cell-associated folate-binding protein is probably produced by de novo protein synthesis, the soluble folate-binding protein seems to be produced from a cellular pool of an already synthesized protein. These results combined with the immunologic cross-reactivity of the two folate-binding proteins strongly suggest a precursor-product relationship between them.

UR - http://www.scopus.com/inward/record.url?scp=0023009070&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023009070&partnerID=8YFLogxK

M3 - Article

VL - 261

SP - 15625

EP - 15631

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 33

ER -