Structure and function of the heat-stable enterotoxin receptor/guanylyl cyclase C

Arc1p is a yeast-specific tRNA-binding protein that forms a ternary complex with glutamyl-tRNA synthetase (GluRSc) and methionyl-tRNA synthetase (MetRS) in the cytoplasm to regulate their catalytic activities and subcellular distributions. Despite Arc1p not being involved in any known biotin-dependent reaction, it is a natural target of biotin modification. Results presented herein show that biotin modification had no obvious effect on the growth-supporting activity, subcellular distribution, tRNA binding, or interactions of Arc1p with GluRSc and MetRS. Nevertheless, biotinylation of Arc1p was temperature dependent; raising the growth temperature from 30 to 37 °C drastically reduced its biotinylation level. As a result, Arc1p purified from a yeast culture that had been grown overnight at 37 °C was essentially biotin free. Non-biotinylated Arc1p was more heat stable, more flexible in structure, and more effective than its biotinylated counterpart in promoting glutamylation activity of the otherwise inactive GluRSc at 37 °C in vitro. Our study suggests that the structure and function of Arc1p can be modulated via biotinylation in response to temperature changes.

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Cell line-specific transcriptional activation of the promoter of the human guanylyl cyclase C/heat-stable enterotoxin receptor gene

Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression ◽

10.1016/0167-4781(95)00190-5 ◽

1996 ◽

Vol 1305 (1-2) ◽

pp. 7-10 ◽

Cited By ~ 13

Author(s):

Elizabeth A. Mann ◽

Mary Lynn Jump ◽

Ralph A. Giannella

Keyword(s):

Cell Line ◽

Guanylyl Cyclase ◽

Transcriptional Activation ◽

Receptor Gene ◽

Guanylyl Cyclase C ◽

Heat Stable

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Differential processing of guanylyl cyclase C along villus-crypt axis of rat small intestine

AJP Cell Physiology ◽

10.1152/ajpcell.1997.272.6.c1995 ◽

1997 ◽

Vol 272 (6) ◽

pp. C1995-C2004 ◽

Cited By ~ 6

Author(s):

L. A. Scheving ◽

K. M. Chong

Keyword(s):

Guanylyl Cyclase ◽

Limited Proteolysis ◽

Structural Heterogeneity ◽

Proteolytic Processing ◽

Enterotoxigenic Escherichia Coli ◽

Guanylyl Cyclase C ◽

Heat Stable ◽

Molecular Events ◽

Cell Fractions ◽

Carboxy Terminal

Many strains of enterotoxigenic Escherichia coli produce a heat-stable peptide enterotoxin (STa) that binds to the intestinal receptor guanylyl cyclase C (GC-C). STa receptors are structurally heterogeneous, but the molecular events causing this heterogeneity remain obscure. We examined the influence of cell position along the villus-crypt axis on STa receptor heterogeneity by fractionating EDTA-dissociated cells that detached in a villus-to-crypt direction. STa affinity labeling experiments revealed that the initially released villus “tip” fraction had four major STa binding proteins (STBPs), with relative molecular weight (M(r)) of 150,000, 135,000, 125,000, and 95,000, that did not react with a GC-C carboxy-terminal antibody. Yet succeeding villus cell fractions had major immunoreactive STBPs with M(r) of 275,000 and 250,000. Limited proteolysis of these larger GC-C isoforms produced 1) smaller STBPs that had M(r) similar to those in the initial villus fraction, 2) a 65,000 M(r) protein GC-C isoform that did not bind STa, and 3) elevated basal and STa-induced cyclase activity. Our data show that STBP structural heterogeneity in the intact intestine arises largely from multisite proteolytic processing of GC-C.

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