scholarly journals Labelling of prolyl hydroxylase tetrameric subunits in freshly isolated chick-embryo tendon cells and in certain chick-embryo tissues in vivo

1982 ◽  
Vol 204 (3) ◽  
pp. 737-742 ◽  
Author(s):  
K Majamaa ◽  
J Oikarinen
Keyword(s):  
Chick Embryo ◽  
Prolyl Hydroxylase ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Beta Subunits ◽  
Chase Period ◽  
Tendon Cells ◽  
Alpha Radioactivity ◽  
A Minor

The labelling of the subunits of prolyl 4-hydroxylase tetramers was studied in freshly isolated chick-embryo tendon cells and in chick-embryo tissues. In the former both the alpha- and beta-subunits of the tetramer were labelled during a 4 h labelling and 2 h chase period, although the radioactivity in the beta-subunit was much lower than in the alpha-subunit. The corresponding subunits of the enzyme from 12-day chick-embryo cartilaginous bone and heart were labelled in 7 h, again the beta-subunit much less than the alpha-subunit, the ratio of radioactivity in the beta-subunit to that in the alpha-subunit (beta/alpha-radioactivity) being 0.20 and 0.32 respectively. The beta/alpha-radioactivity then increased almost linearily with time between 7 and 24 h, by 9.5-fold in the cartilaginous bone and 3-fold in the heart, and beta/alpha-radioactivity values above 1.0 were reached. The free beta-subunit-size protein (the beta'-protein), which is also present in cells, had been labelled quite heavily by 7 h. The beta/alpha-radioactivity at 7h, determined in four tissues with different ratios of prolyl hydroxylase tetramers to total immunoreactive protein (tetramer percentage), was low in tissues with a high tetramer percentage. It is thus proposed that only a minor fraction of the beta'-protein must be processed to the tetrameric beta-subunit and utilized in the synthesis of the prolyl 4-hydroxylase tetramers.

scholarly journals Investigations into the post-translational modification and mechanism of isopenicillin N:acyl-CoA acyltransferase using electrospray mass spectrometry

1993 ◽  
Vol 294 (2) ◽  
pp. 357-363 ◽  
Author(s):  
R T Aplin ◽  
J E Baldwin ◽  
P L Roach ◽  
C V Robinson ◽  
C J Schofield
Keyword(s):  
Mass Spectrometry ◽  
Electrospray Mass Spectrometry ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
British Library ◽  
Beta Subunits ◽  
Post Translational Modification ◽  
Acetic Acids

Electrospray mass spectrometry (e.s.m.s.) was used to confirm the position of the post-translational cleavage of the isopenicillin N:acyl-CoA acyltransferase preprotein to give the alpha- and beta-subunits. The e.s.m.s. studies suggested partial modification of the alpha-subunit in vivo by exogenously added substituted acetic acids. E.s.m.s. has also allowed the observation in vitro of the transfer of the acyl group from several acyl-CoAs to the beta-subunit. N.m.r. data for the CoA species have been deposited as Supplementary Publication SUP 500173 (2 pages) at the British Library Document Supply Centre (DSC), Boston Spa, Wetherby, West Yorkshire LS23 7BQ, from whom copies can be obtained on the terms indicated in Biochem. J. (1993) 289, 9.

Secretion of alpha and TSH-beta subunits in patients with acromegaly: an in vivo study

1990 ◽  
Vol 122 (6) ◽  
pp. 729-734
Author(s):  
Hossein Assadian ◽  
Akira Shimatsu ◽  
Hiroyuki Koshiyama ◽  
Naoki Hattori ◽  
Yasuhiro Ishikawa ◽  
...  
Keyword(s):  
Pituitary Adenomas ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
In Vivo Study ◽  
Beta Subunits ◽  
Active Acromegaly

Abstract. Plasma alpha and TSH-beta subunit responses to iv administration of GHRH were examined in 19 patients with active acromegaly. In 4 patients (21%), plasma alpha subunit levels were increased over 50% of basal levels after administration of GHRH, whereas plasma TSH-beta subunit levels were increased in response to GHRH in another 5 patients (26%). No patient showed simultaneous increases of alpha and beta subunits. After successful surgery, alpha and TSH-beta subunits did not respond to GHRH. These findings support the idea that some pituitary adenomas in acromegaly co-secrete GH and either alpha subunit or TSH-beta subunit.

A spectrin-like protein from mouse brain membranes: phosphorylation of the 235,000-dalton subunit

1984 ◽  
Vol 247 (1) ◽  
pp. C61-C73 ◽  
Author(s):  
S. R. Goodman ◽  
I. S. Zagon ◽  
C. F. Whitfield ◽  
L. A. Casoria ◽  
S. B. Shohet ◽  
...  
Keyword(s):  
Mouse Brain ◽  
Beta Subunit ◽  
Beta Subunits ◽  
Mouse Erythrocyte ◽  
Alpha Beta ◽  
Beta 2 ◽  
Independent Protein ◽  
Brain Spectrin

A mouse brain spectrin-like protein, which was an immunoreactive analogue of erythrocyte spectrin, has been isolated from demyelinated membranes. This spectrin analogue was a 10.5 S, 972,000 molecular weight (Mr) (alpha beta)2 tetramer containing subunits of 240,000 (alpha) and 235,000 (beta) Mr. We demonstrated that in vivo only the 235,000 Mr beta subunit of the mouse brain spectrin-like protein was phosphorylated, which was an analogous situation to mouse erythrocyte spectrin in which only the 220,000 Mr beta subunit was phosphorylated. Incubation of isolated membrane fractions with [gamma-32P]ATP +/- adenosine 3',5'-cyclic monophosphate (cAMP) indicated that mouse brain spectrin-like protein, mouse erythrocyte spectrin, and human erythrocyte spectrin's beta subunits were all phosphorylated in vitro by membrane-associated cAMP-independent protein kinases.

scholarly journals Effect of cortisol acetate on collagen biosynthesis and on the activities of prolyl hydroxylase, lysyl hydroxylase, collagen galactosyltransferase and collagen glucosyltransferase in chick-embryo tendon cells

1977 ◽  
Vol 164 (3) ◽  
pp. 533-539 ◽  
Author(s):  
A Oikarinen
Keyword(s):  
Chick Embryo ◽  
Enzyme Activities ◽  
Collagen Synthesis ◽  
Enzyme Synthesis ◽  
Prolyl Hydroxylase ◽  
Chick Embryos ◽  
Isolated Cells ◽  
Lysyl Hydroxylase ◽  
Tendon Cells

Collagen synthesis and the activities of prolyl hydroxylase, lysyl hydroxylase, collagen galactosyltransferase and collagen glucosyltransferase were studied in isolated chick-embryo tendon cells after the administration of cortisol acetate to the chick embryos. When the steroid was injected 1 day before isolation of the tendon cells, collagen synthesis was decreased, even though the enzyme activities were not changed. When cortisol acetate was given as repeated injections over a period of 4 days, both collagen synthesis and the enzyme activities decreased. The hydroxylase activities decreased even more than the two collagen glycosyltransferase activities, both in isolated cells and in whole chick embryos. The amount of prolyl hydroxylase protein diminished to the same extent as the enzyme activity, indicating that cortisol acetate inhibits enzyme synthesis. The inhibitory effect of cortisol acetate on collagen synthesis and on the enzyme activities was partially reversible in 3 days. Total protein synthesis was completely restored within this time. Only massive doses of cortisol acetate inhibited collagen synthesis in vitro. Additional experiments indicated that cortisol acetate did not decrease the rate of the enzyme reactions when added directly to the enzyme incubation mixtures. The results suggest that cortisol acetate decreases collagen synthesis both by its direct effect on collagen polypeptide-chain synthesis and by decreasing the activities of enzymes involved in post-translational modifications.

Analysis of subunit assembly of the Na-K-ATPase

1994 ◽  
Vol 266 (3) ◽  
pp. C579-C589 ◽  
Author(s):  
D. M. Fambrough ◽  
M. V. Lemas ◽  
M. Hamrick ◽  
M. Emerick ◽  
K. J. Renaud ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Sodium Pump ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Beta Subunits ◽  
Limited Region ◽  
Subunit Assembly ◽  
Alpha Subunits ◽  
Subunit Isoforms ◽  
P Type

The Na-K-ATPase, or sodium pump, is comprised of two subunits, alpha and beta. Each subunit spans the lipid bilayer of the cell membrane. This review summarizes our efforts to determine how the two subunits interact to form the functional ion transporter. Our major approach has been to observe the potential for subunit assembly when one or both subunits are truncated or present as chimeras that retain only a limited region of the Na-K-ATPase. DNAs encoding these altered subunit forms of the avian Na-K-ATPase are expressed in mammalian cells. Monoclonal antibodies specific for the avian beta-subunit are then used to purify newly synthesized avian beta-subunits, and the presence of accompanying alpha-subunits indicates that subunit assembly has occurred. The ectodomain of the beta-subunit (approximately residues 62-304) is sufficient for assembly with the alpha-subunit, and a COOH-terminal truncation of the beta-subunit that lacks aminoacyl residues beyond 162 will assemble inefficiently. A maximum of 26 aminoacyl residues of the alpha-subunit are necessary for robust assembly with the beta-subunit, when this sequence replaces the COOH-terminal half of the loop between membrane spans 7 and 8 in the SERCA1 Ca-ATPase. This region of the Ca-ATPase faces the lumen of the endoplasmic reticulum. These findings encourage study of other related questions, including whether there is preferential assembly of certain subunit isoforms and how various P-type ATPases are targeted to their appropriate subcellular compartments.

Free α-Subunit, Free β-Subunit of Human Chorionic Gonadotropin (hCG), and Intact hCG in Sera of Healthy Individuals and Testicular Cancer Patients

1992 ◽  
Vol 38 (3) ◽  
pp. 370-376 ◽  
Author(s):  
S Madersbacher ◽  
R Klieber ◽  
K Mann ◽  
C Marth ◽  
M Tabarelli ◽  
...  
Keyword(s):  
Testicular Cancer ◽  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Serum Concentrations ◽  
Healthy Individuals ◽  
Pathological Conditions

Abstract To determine the serum concentrations of human chorionic gonadotropin (hCG), its free beta-subunit (hCG beta), and the free alpha-subunit (free alpha) common to all human glycoprotein hormones under physiological and pathological conditions, we developed monoclonal antibody-based immunoenzymometric assays. Free alpha-subunit was detected in the sera of all healthy individuals of both sexes; hCG was measurable in sera of 54% of the men, and 46% were positive for free hCG beta; in nonpregnant women, 69.5% were positive for hCG, 68.4% for the free beta-subunit. Pathological conditions, i.e., hCG-producing tumors, were studied in vitro and in vivo. In vitro, the concentrations of hCG, free hCG beta, and free alpha in tissue-culture supernates of a choriocarcinoma cell-line ("JAR") showed a parallel pattern during time-course analysis. In vivo, in long-term follow-up studies of 13 patients with testicular cancer, serum concentrations of the three analytes paralleled each other, whether the disease was in remission or not. Because of a selective increase of free hCG beta and free alpha in 27% of seminomatous tumor patients and in 13% of the nonseminomatous patients, the percentage of tumor-marker-positive sera was increased from 15% to 42% and 57% to 70%, respectively, by the additional measurement of free hCG beta and free alpha. Thus hCG, free hCG beta, and free alpha are physiologically present in a high percentage of the sera from healthy men, and the determination of free hCG beta and free alpha, although not of prognostic value, improves the diagnostic possibilities in patients with testicular cancer.

Na(+)-K(+)-ATPase alpha 1- and beta 1-subunit degradation: evidence for multiple subunit specific rates

1993 ◽  
Vol 264 (3) ◽  
pp. C583-C590 ◽  
Author(s):  
L. Lescale-Matys ◽  
D. S. Putnam ◽  
A. A. McDonough
Keyword(s):  
Beta Subunit ◽  
Beta Subunits ◽  
Plasma Membrane Protein ◽  
Significant Component ◽  
Chase Period ◽  
Pig Kidney ◽  
Alpha Subunits ◽  
Degradation Rates ◽  
Synthesis And Degradation ◽  
Rate Of Accumulation

Na(+)-K(+)-ATPase is a heterodimeric plasma membrane protein consisting of an alpha-catalytic and a beta-glycoprotein subunit. Because these two subunits are derived from two separate genes, they may not be synthesized with stoichiometric equivalence. The aim of this study was to estimate relative rates of synthesis and degradation of nascent and mature Na(+)-K(+)-ATPase alpha- and beta-subunits to determine whether either of the nascent subunits accumulates in excess and, if so, the fate of the excess subunits. We studied a pig kidney cell line (LLC-PK1/Cl4) that expresses only alpha 1- and beta 1-subunits. Relative synthesis and degradation rates of nascent subunits were first estimated by pulsing cells for 10 min with [35S]methionine followed by chase periods of up to 120 min and by immunoprecipitation. We found that directly after labeling, beta-subunits were present in threefold excess over alpha-subunits and that nearly 50% of this beta-subunit pool was degraded by 60 min. Nascent alpha-subunits were not degraded during the chase period. In a second strategy to examine relative rates of nascent alpha- vs. beta-subunit accumulation, cells were pulsed for 5-60 min and immunoprecipitated directly (without chase). The rate of accumulation of labeled alpha was greater than that of beta between 5 and 60 min, consistent with the results of the pulse-chase strategy, demonstrating a significant component of degradation of beta during this period. Despite the very different degradation rates of newly synthesized alpha- vs. beta-subunits, the degradation rates of alpha- and beta-subunits beyond 4 h after synthesis were indistinguishable (t0.5 = 10-12 h).(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals A human leukocyte differentiation antigen family with distinct alpha-subunits and a common beta-subunit: the lymphocyte function-associated antigen (LFA-1), the C3bi complement receptor (OKM1/Mac-1), and the p150,95 molecule.

1983 ◽  
Vol 158 (6) ◽  
pp. 1785-1803 ◽  
Author(s):  
F Sanchez-Madrid ◽  
J A Nagy ◽  
E Robbins ◽  
P Simon ◽  
T A Springer
Keyword(s):  
Human Leukocyte ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Beta Subunits ◽  
Complement Receptor ◽  
Lymphocyte Function ◽  
Subunit Dissociation ◽  
Molecular Weights ◽  
Alpha Subunits ◽  
Sds Page

The human lymphocyte function-associated antigen-1 (LFA-1), the complement receptor-associated OKM1 molecule, and a previously undescribed molecule termed p150,95, have been found to be structurally and antigenically related. Each antigen contains an alpha- and beta-subunit noncovalently associated in an alpha 1 beta 1-structure as shown by cross-linking experiments. LFA-1, OKM1, and p150,95 alpha-subunit designations and their molecular weights are alpha L = 177,000 Mr, alpha M = 165,000 Mr, and alpha X = 150,000 Mr, respectively. The beta-subunits are all = 95,000 Mr. Some MAb precipitated only LFA-1, others only OKM1, and another precipitates all three antigens. The specificity of these MAb for particular subunits was examined after subunit dissociation by high pH. MAb specific for LFA-1 or OKM1 bind to the alpha L- or alpha M-subunits, respectively, while the cross-reactive MAb binds to the beta-subunits. Coprecipitation experiments with intact alpha 1 beta 1-complexes showed anti-alpha and anti-beta MAb can precipitate the same molecules. In two-dimensional (2D) isoelectric focusing-SDS-PAGE, the alpha subunits of the three antigens are distinct, while the beta-subunits are identical. Biosynthesis experiments showed alpha L, alpha M, and alpha X are synthesized from distinct precursors, as is beta. The three antigens differ in expression on lymphocytes, granulocytes, and monocytes. During maturation of the monoblast-like U937 line, alpha M and alpha X are upregulated and alpha L is downregulated. Some MAb to the alpha subunit of OKM1 inhibited the complement receptor type three. LFA-1, OKM1, and p150,95 constitute a novel family of functionally important human leukocyte antigens that share a common beta-subunit.

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