The interaction of an anionic photoreactive probe with the anion transport system of the human red blood cell

Cabantchik, Z. I.; Knauf, Philip A.; Ostwald, Thomas J.; Markus, H.; Davidson, Lorinda; Breuer, William; Rothstein, Aser

doi:10.1016/0005-2736(76)90322-9

Cited by 61 publications

(36 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The basis of selectivity is not size alone, for bulkier ions are transported, though slowly; examples are tetrathionate (Deuticke et al, 1978), benzenesulfonate, phthalate, p-aminobenzenesulfonate (Aubert & Motais, 1975), pyridoxal phosphate (Nanri, Hamasaki & Minakami, 1983), and even NAP-taurine (Cabantchik et al, 1976;Rothstein, Knauf & Cabantchik, 1977;Knauf et al, 1978a). Hence the breadth of the channel through which the substrate is conducted would not seem to determine the specificity for ions no larger than sulfate or phosphate.…”

Section: The Expression Of Substrate Specificitymentioning

confidence: 95%

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

1989

J. Membrain Biol.

View full text Add to dashboard Cite

The transition-state theory of exchange-only membrane transport is applied to experimental results in the literature on the anion exchanger of red cells. Two central features of the system are in accord with the theory: (i) forming the transition state in translocation involves a carrier conformational change; (ii) substrate specificity is expressed in transport rates rather than affinities. The expression of specificity is consistent with other evidence for a conformational intermediate (not the transition state) formed in the translocation of all substrates. The theory, in conjunction with concepts derived from the chemistry of macrocyclic ion inclusion complexes, prescribes certain essential properties in the transport site. Separate subsites are required for the preferred substrates, Cl- and HCO3-, to account for tight binding in the transition state (Kdiss congruent to 1 microM). Further, the following mechanism is suggested. A substrate anion initially forms a loose surface complex at one subsite, but in the transition state the subsites converge to form an inclusion complex in which the binding forces are greatly increased through a chelation effect. The conformational change at the substrate site, which is driven by the mounting forces of binding, sets in train a wider conformational change that converts the carrier from an immobile to a mobile form. Though simple, this composite-site mechanism explains many unusual features of the system. It accounts for substrate inhibition, partially noncompetitive inhibition of one substrate by another, and "tunneling," which is net transport under conditions where exchange should prevail, according to other models. All three types of behavior result from the formation of a ternary complex in which substrate anions are bound at both subsites. The mechanism also accounts for the enormous range of substrate structures accepted by the system, for the complex inhibition by the organic sulfate NAP-taurine, and for the involvement of several cationic side chains and two different protein domains in the transport site.

show abstract

Section: The Expression Of Substrate Specificitymentioning

confidence: 95%

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

1989

J. Membrain Biol.

View full text Add to dashboard Cite

show abstract

“…One additional agent, NAP-taurine, which inhibits SO4 fluxes in erythrocytes [4] and hepatocytes [7], was tested in gluconate Ringer's containing 0.22 mM SO4 and 20 mM C1. The experiment was performed in the dark since photolysis converts NAP-taurine to a highly reactive nitrene which acts as a general covalent labeling reagent.…”

Section: Effects Of Inhibitorsmentioning

confidence: 99%

Sulfate transport in rabbit ileum: Characterization of the serosal border anion exchange process

Langridge-Smith

Field

1981

J. Membrain Biol.

View full text Add to dashboard Cite

The preceding paper [30] shows that transepithelial ileal SO4 transport involves Na-dependent uptake across the ileal brush border, and Cl-dependent efflux across the serosal border. The present study examines more closely the serosal efflux process. Transepithelial mucosa (m)-to-serosa (s) and sto-m fluxes (Jms, Jsm) across rabbit ileal mucosa were determined under short-circuit conditions. SO4 was present at 0,22 mM. In standard C1, HCO3 Ringer's, jso4 was 81.3 +5.3 (1SE) and jso4 was 2.5_+0.2 nmol cm-2hr-l(n=20). Serosal addition of 4-acetamido-4'-isothiocyanostilbene-22'-disulfonate (SITS), 44'-diisothiocyanostilbene-22'-disulfonate (DIDS) or 1-anilino-8-naphthalene-sulfonate (ANS) inhibited SO4 transport, SITS being the most potent. Several other inhibitors of anion exchange in erythrocytes and other cells had no effect on ileal SO4 fluxes. In contrast to its effect on SO4 transport, SITS (500 gM) did not detectably alter C1 transport.Replacement of all C1, HCO3 and PO~ with gluconate reduced jso4 by 70% and increased jso4 by 400%. A small but significant I s~ remained, lso4 vnet ~ms could be increased by addition to the serosal side of C1, Br, I, NO3 or SO4. The stimulatory effect of all these anions was saturable and SITS-inhibitable. The maximal jso4 in the presence of C1 was considerably higher than in the presence of SO4 (73.1 and 42.2nmol cm -2 hr -t, respectively; p<0.001). The K~ value for C1 was 7.4 mM, 10-fold higher than that for SO4 (0.7 mM). Omitting HCO3 and PO4 had no measurable effects on SO4 fluxes.This study shows that (i) SO4 crosses the serosal border of rabbit ileal mucosa by anion exchange; (i 0 the exchange process is inhibited by SITS, DIDS and ANS, but not by several other inhibitors of anion exchange in other systems; (iii) SO4 may exchange for C1, Br, I, NO3 and SO4 itself, but probably not for HCO3 or PO~; (iv) kinetics of the exchange system suggest there is a greater affinity for SO4 than for C1, although the maximal rate of exchange is higher in the presence of C1; and, finally (v) SITS has little or no effect on net C1 transport.The preceding paper [30] shows that active SO4 absorption in rabbit ileum involves two separate steps: (0 a Na-dependent uptake at the brush border, and (ii) a Cl-dependent efflux at the serosal border. The latter can be blocked by the stilbene derivatives 4-acetamido-4'-isothiocyanostilbene-2,T-disulfonate (SITS) 1 and 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS), which are potent inhibitors of anion exchange in both erythrocytes [5] and Ehrlich ascites tumor cells [22,23]. In the present study, we further investigate the serosal border component of SO4 transport in rabbit ileum. Transepithelial mucosa (m)-to-serosa (s) and s-to-m fluxes of SO4 were determined under short-circuit conditions in Ringer's solutions of different anionic compositions. The effects of a number of anion transport inhibitors were also tested. For most flux measurements, the SO~ concentration was set at 0.22 mM, or about 10-fold lower than in the preceding study [30], ...

show abstract

“…In recent years, our understanding of the anion exchange system of the human red cell has been greatly augmented by two sorts of information. On the one hand, kinetic studies have shown that the transport is almost exclusively a onefor-one exchange of anions and that the system displays properties consistent with a mobile carrier model (Wieth, 1972;Gunn et al, 1973;Hunter, 1971; Sachs et al, 1975;Cabantchik et al?). On the other hand, the use of inhibitory chemical probes has led to the identification of the 95,000dalton polypeptide, known as band 3 (Fairbanks et al, 1971), as the protein which apparently mediates the transport process (Cabantchik and Rothstein, 1974;Ho and Guidotti, 1975;Rothstein et al, 1976).…”

Section: Introductionmentioning

confidence: 99%

“…To integrate these two kinds of information, additional knowledge is required concerning the relationship between carrier sites postulated from kinetic analysis of transport and inhibitory sites in band 3 labeled by the covalent reactions of the chemical probes. The evaluation of the mechanism by which probes inhibit transport can be greatly facilitated by the use of "bimodal" inhibitors (Deuticke, 1977;Cabantchik et al)), that is, probes which are capable of covalent reaction for labeling purposes but which, under other circumstances, act as reversible inhibitors of the system, allowing the nature of their effects to be determined by a kinetic analysis. Photoaffinity reagents are particularly useful, because they can be used as reversible or irreversible probes under exactly the same conditions, the only difference being the presence or absence of light to activate the compound.…”

Section: Introductionmentioning

confidence: 99%

Asymmetry of the red cell anion exchange system. Different mechanisms of reversible inhibition by N-(4-azido-2-nitrophenyl)-2-aminoethylsulfonate (NAP-taurine) at the inside and outside of the membrane.

Knauf¹,

Ship²,

Breuer³

et al. 1978

The Journal of General Physiology

View full text Add to dashboard Cite

In the dark, the photoaffinity reagent, N-(4-azido-2-nitrophenyl)-2-aminoethylsulfonate (NAP-taurine), acts as a reversible inhibitor of red cell anion exchange when it is present either within the cell or in the external solution. A detailed analysis of the inhibition kinetics, however, reveals substantial differences in the responses to the probe at the two sides of the membrane. On the inside of the cell, NAP-taurine is a relatively low affinity inhibitor of chloride exchange (Ki = 370 microM). Both the effects of chloride on NAP-taurine inhibition and the affinity of NAP-taurine for the system as a substrate are consistent with the concept that internal NAP-taurine competes with chloride for the substrate site of the anion exchange system. External NAP-taurine, on the other hand, is a far more potent inhibitor of chloride exchange (Ki = 20 microM). It acts at a site of considerably lower affinity for chloride than the substrate site, probably the modifier site, at which halide anions are reported to cause a noncompetitive inhibition of chloride transport. NAP-taurine therefore seems to interact preferentially with either the substrate or modifier site of the transport system, depending on the side of the membrane at which it is present. It is suggested that the modifier site is accessible to NAP-taurine only from the outside whereas the transport site may be accessible from either side.

show abstract

The interaction of an anionic photoreactive probe with the anion transport system of the human red blood cell

Cited by 61 publications

References 39 publications

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

Role of substrate binding forces in exchange-only transport systems: II. Implications for the mechanism of the anion exchanger of red cells

Sulfate transport in rabbit ileum: Characterization of the serosal border anion exchange process

Asymmetry of the red cell anion exchange system. Different mechanisms of reversible inhibition by N-(4-azido-2-nitrophenyl)-2-aminoethylsulfonate (NAP-taurine) at the inside and outside of the membrane.

Contact Info

Product

Resources

About