scholarly journals The Na-K-Cl cotransport protein of shark rectal gland. I. Development of monoclonal antibodies, immunoaffinity purification, and partial biochemical characterization.

1992 ◽  
Vol 267 (35) ◽  
pp. 25428-25437 ◽  
Author(s):  
C Lytle ◽  
J.C. Xu ◽  
D Biemesderfer ◽  
M Haas ◽  
B Forbush
Keyword(s):  
Monoclonal Antibodies ◽  
Biochemical Characterization ◽  
Rectal Gland ◽  
Immunoaffinity Purification ◽  
Shark Rectal Gland

Cyclic AMP stimulates ouabain-insensitive ion movement in shark rectal gland

1984 ◽  
Vol 154 (2) ◽  
pp. 139-144 ◽  
Author(s):  
Patricio Silva ◽  
Kate Spokes ◽  
Jonathan A. Epstein ◽  
Arthur Stevens ◽  
Franklin H. Epstein
Keyword(s):  
Cyclic Amp ◽  
Rectal Gland ◽  
Ion Movement ◽  
Shark Rectal Gland

scholarly journals Characterization of four new monoclonal antibodies against the distal N-terminal region of PrPc

2014 ◽  
Author(s):  
Alessandro Didonna ◽  
Anja Colja Venturini ◽  
Katrina Hartman ◽  
Tanja Vranac ◽  
Vladka Curin Serbec ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Prion Protein ◽  
Biochemical Characterization ◽  
Prion Diseases ◽  
Physiological Role ◽  
Prion Infection ◽  
Promising Tool ◽  
C Terminus ◽  
N Terminus

Prion diseases are a group of fatal neurodegenerative disorders that affect humans and animals. They are characterized by the accumulation in the central nervous system of a pathological form of the host-encoded prion protein (PrPC). The prion protein is a membrane glycoprotein that consists of two domains: a globular, structured C-terminus and an unstructured N-terminus. The N-terminal part of the protein is involved in different functions in both health and disease. In the present work we discuss the production and biochemical characterization of a panel of four monoclonal antibodies (mAbs) against the distal N-terminus of PrPC using a well-established methodology based on the immunization of Prnp0/0 mice. Additionally, we show their ability to block prion (PrPSc) replication at nanomolar concentrations in a cell culture model of prion infection. These mAbs represent a promising tool for prion diagnostics and for studying the physiological role of the N-terminal domain of PrPC.

cAMP-activated C1 conductance is expressed in Xenopus oocytes by injection of shark rectal gland mRNA

1991 ◽  
Vol 260 (3) ◽  
pp. C664-C669 ◽  
Author(s):  
S. K. Sullivan ◽  
K. Swamy ◽  
M. Field
Keyword(s):  
Xenopus Oocytes ◽  
Functional Characterization ◽  
Equilibrium Potential ◽  
Rectal Gland ◽  
Calcium Buffer ◽  
Cl Channel ◽  
Shark Rectal Gland ◽  
Cl Channels ◽  
Cl Conductance

Development of reliable expression systems for use in identification and functional characterization of proteins required for secretory Cl channel activity is key to understanding the molecular basis of cystic fibrosis (CF). Until now, heterologous expression of epithelial Cl channels had not been accomplished. We show here that Xenopus oocytes express an adenosine 3',5'-cyclic monophosphate (cAMP)-activated Cl conductance after injection of mRNA from shark rectal gland. Current through this conductance was rapidly activated by intracellular application of cAMP, reversed near the chloride equilibrium potential (ECl), blocked by the Cl channel inhibitor 5-nitro-2-(3-phenylpropylamino) benzoate, and was not affected by preincubation with the intracellular calcium buffer bis-(2-amino-5-methylphenoxy)-ethane-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester, a condition that prohibits activation of the endogenous Ca-activated Cl conductance.

Immunologic analysis of human breast cancer progesterone receptors. 1. Immunoaffinity purification of transformed receptors and production of monoclonal antibodies

Biochemistry ◽  
1987 ◽  
Vol 26 (19) ◽  
pp. 6250-6262 ◽  
Author(s):  
Patricia A. Estes ◽  
Eric J. Suba ◽  
Janet Lawler-Heavner ◽  
Dorraya El-Ashry-Stowers ◽  
Lisa L. Wei ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Monoclonal Antibodies ◽  
Human Breast Cancer ◽  
Progesterone Receptors ◽  
Human Breast ◽  
Immunoaffinity Purification ◽  
Immunologic Analysis

Cl- secretion by cultured shark rectal gland cells. II. Effects of forskolin on cellular electrophysiology

1991 ◽  
Vol 260 (4) ◽  
pp. C824-C831 ◽  
Author(s):  
W. M. Moran ◽  
J. D. Valentich
Keyword(s):  
Apical Membrane ◽  
Basolateral Membrane ◽  
Electrical Potential ◽  
Ringer Solution ◽  
Rectal Gland ◽  
Electrical Potential Difference ◽  
Electrophysiological Properties ◽  
Shark Rectal Gland ◽  
Membrane Electrical Potential ◽  
Cl Conductance

Employing microelectrode techniques we have assessed the cellular electrophysiological properties of shark rectal gland (SRG) cells in primary culture. In the absence of secretagogues a 10-fold reduction in the Cl- concentration of the apical superfusate shark Ringer solution had little effect on either apical membrane electrical potential difference (Va) or fractional resistance (fRa), indicating little, if any, apical membrane Cl- conductance. Superfusing the basolateral surface with high-K+ shark Ringer solution (K+ increased 10-fold) depolarized the basolateral membrane electrical potential difference (Vb) by 43 mV, indicating that this barrier is largely K+ conductive. In addition, basolateral Ba2+ (5 mM) depolarized Vb by 12 mV and reduced fRa from 0.92 to 0.58, results consistent with a K(+)-conductive basolateral membrane in unstimulated SRG cells. Basolateral forskolin (10(-6) M) depolarized Va by 25 mV and caused a dramatic reduction in fRa from 0.97 to approximately 0.10. Under these conditions, a 10-fold decrease in apical superfusate Cl- concentration depolarized Va by 37 mV, revealing an adenosine 3',5'-cyclic monophosphate-induced apical membrane Cl- conductance. The time course of the forskolin-induced changes in Va and Vb suggests that the basolateral membrane K+ conductance increased and maintained the driving force for apical Cl- exit, as in other Cl(-)-secreting epithelia. These electrophysiological properties compare favorably with those of the perfused SRG tubule and indicate that SRG primary cultures are a suitable model for Cl(-)-secreting epithelia.

Propionate induces cell swelling and K+ accumulation in shark rectal gland

1989 ◽  
Vol 257 (2) ◽  
pp. C377-C384 ◽  
Author(s):  
G. M. Feldman ◽  
F. N. Ziyadeh ◽  
J. W. Mills ◽  
G. W. Booz ◽  
A. Kleinzeller
Keyword(s):  
Atpase Activity ◽  
Propionic Acid ◽  
Exchange Process ◽  
Squalus Acanthias ◽  
Cell Swelling ◽  
Rectal Gland ◽  
Structural Elements ◽  
Acid Diffusion ◽  
Shark Rectal Gland ◽  
Solute Accumulation

Small organic anions have been reported to induce cell solute accumulation and swelling. To investigate the mechanism of swelling, we utilized preparations of rectal gland cells from Squalus acanthias incubated in medium containing propionate. Propionate causes cells to swell by diffusing across membranes in its nonionic form, acidifying cell contents, and activating the Na+-H+ antiporter. The Na+-H+ exchange process tends to correct intracellular pH (pHi), and thus it maintains a favorable gradient for propionic acid diffusion and allows propionate to accumulate. Activation of the Na+-H+ antiport also facilitates Na+ entry into the cell and Nai accumulation. At the same time Na+-K+-ATPase activity, unaffected by propionate, replaces Nai with Ki, whereas the K+ leak rate, decreased by propionate, allows Ki to accumulate. As judged by 86Rb+ efflux, the reduction in K+ leak was not due to propionate-induced cell acidification or reduction in Cli concentration. Despite inducing cell swelling, propionate did not disrupt cell structural elements and F actin distribution along cell membranes.

Elucidation of the Binding Regions of PAI-1 Neutralizing Antibodies Using Chimeric Variants of Human and Rat PAI-1

2001 ◽  
Vol 85 (05) ◽  
pp. 866-874 ◽  
Author(s):  
A. P. Bijnens ◽  
T. H. Ngo ◽  
A. Gils ◽  
J. Dewaele ◽  
I. Knockaert ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Plasminogen Activator ◽  
Neutralizing Antibodies ◽  
Rational Design ◽  
Biochemical Characterization ◽  
Tissue Type ◽  
Binding Region ◽  
Plasminogen Activator Inhibitor 1 ◽  
Inhibitory Antibodies ◽  
Pai 1

SummaryIncreased levels of plasminogen activator inhibitor-1 (PAI-1), the main physiological inhibitor of tissue-type plasminogen activator (t-PA) in plasma, are a known risk factor for thromboembolic and cardiovascular diseases. The elucidation of the binding site of inhibitory monoclonal antibodies may contribute to the rational design of PAI-1 modulating therapeutics. In this study, homolog-scanning mutagenesis was used to identify the binding region of a variety of human PAI-1 inhibitory antibodies, lacking cross-reactivity with rat PAI-1. Therefore, eight chimeric human/rat PAI-1 variants, containing rat PAI-1 substitutions at the N-terminal or C-terminal end with splicing sites at positions 26, 81, 187, 277 or 327, were generated and purified. Biochemical characterization revealed that all chimeras were folded properly. Subsequently, surface plasmon resonance was used to determine the affinity of various monoclonal antibodies for these chimera. Comparative analysis of the affinity and ELISA data allowed the identification of the major binding region of the inhibitory antibodies MA-8H9D4, MA-33B8F7, MA-44E4, MA-42A2F6 and MA-56A7C10. Thus, three segments in human PAI-1 containing each at least one site involved in the neutralization of PAI-1 activity could be identified, i.e. (1) the segment from residue 81 to residue 187 (comprising -helices hD, hE and hF, -strands s4C, s3A, s2A and s1A and the loops connecting these elements), (2) the segment between residues 277 and 327 (hI, thIs5A, s5A and s6A) and (3) the region C-terminal from amino acid 327, including the reactive site loop. The current data, together with previous data, indicate that PAI-1 contains at least four different regions that could be considered as putative targets to modulate its activity.

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