Abstract
Characterization of the Na+/K+ pump was by no means completed in the 1950s. Essential concerns—from assessing conflicting reports to generalizing isolated claims—awaited resolution. But beyond clarifications and elaborations lay a basic question: What is the Na+/K+ pump? The reductionistic program of biological explanation translates that query as: What is the chemical nature of the pump? A related question is: How does this chemical nature endow it with the functional properties observed?
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Notes to Chapter 9
Glynn (1957b), p. 297.
Ibid., p. 295.
Ibid., pp. 296, 297.
Clarkson and Maizels (1952). They discussed the activity as an “apyrase,” meaning that the final product of ATP hydrolysis was AMP; by contrast, ADP is the final product with an ATPase. But an ATPase in the presence of adenylate kinase will also produce AMP from ATP. With hindsight I refer to ATPase activity.
Herbert (1956), p. 29. The Na concentration equivalent to “0.4%” is 150 mM. He does not say whether or not K was present.
Dunham (1957b).
The attempt is feeble because many enzymes have similar pH optima: it is not a discriminating characteristic.
Skou (1957a).
Ibid., p. 394. Skou’s introduction implies that he undertook the experiments expressly to study cation transport. Skou later acknowledged that this presentation was a “subsequent rationalization” ( 1989, p. 436 ).
ATP is an acid, and neutralization with a base necessarily introduces some cation. Skou converted ATP (obtainable commercially as some salt) to the free acid-by passing it through an ion exchange column in the H+ form-and then neutralized it with the organic base 2-amino-2-methyl-1,3-propanediol.
I refer to the ATPase activity of impure enzyme preparations due to Na’ and K+ as “(Na’ + K+)-stimulated ATPase activity”; I reserve “Na+/K+-ATPase” for the actual enzyme.
Skou (1957a), p. 401.
This account is based on Skou (1989), Post (1974, 1989), interviews and correspondence with Skou, Post, Hoffman, Tosteson, and Nerby (1993–1995), and Skou’s examination of his laboratory notebooks (1993).
Libet (1954).
Abood and Gerard (1954).
Through 1964, Skou’s 13 papers on this subject were cited nearly 150 times.
Skou’s laboratory notebook for 28 June 1955 (the last workday before he left for vacation) asks: “ATP-is it Na+ or K+?”. The reminiscences (Skou, 1989, p. 436, and Post, 1974, p. 6) imply, misleadingly, that the time spent untangling the sources of the erratic results was far longer.
Skou attributed the delay in publishing to his primary interest continuing to be local anesthetics, to his unfamiliarity with English, and to the absence-in his environment-of a sense of urgency, either from a fear of competitors or from a need to build a bibliography.
Utter (1950). Utter presented his results as apyrase activity, although he explicitly noted that the activity could result from an ATPase plus adenylate kinase.
Steinbach (1954). He also had been studying ATPases, but with Cat+ as the divalent cation (Steinbach, 1949 ).
Glynn (1956, 1957b, 1959).
I have omited a few papers on bacteria, which are irrelevant to this story.
Muntz and Hurwitz (1951).
Novikoff et al. (1952), p. 160. Stimulation by even optimal concentrations of Na+ and K+ would not be large with liver homogenates; Emmelot and Bos (1962) found only 30% stimulation using a plasma membrane fraction.
Gore (1951).
With 5 mM ATP, the Na+ concentration at pH 7.4 would be 15–20 mM. Since K+ was being varied, it presumably was not also present with the glycylglycine (Na+ and K+ were then the standard cations for buffers). At pH 7.4, roughly 15 mM Na+ would be present with 100 mM glycylglycine.
Mcllwain (1952a,b).
Findlay et al. (1954).
Lowry et al. (1954), p. 28.
Skou (1957b). His description was during the discussion period following a paper by Keynes. By the time of the meeting, Skou had already submitted his paper.
Others present included Lowry, Mcllwain, Steinbach, Tosteson, and Ussing.
According to Pope (telephone conversation, 1993), they had previously found effects of cations on ATPase activity but had been interpreting their results differently.
Hess and Pope (1957). “FASEB” is the abbreviation for the Federation of American Societies for Experimental Biology, which held large meetings each spring.
Hess (1962).
Post (1974).
Skou (1989), p. 437. Skou states his supposition that Post had not yet read his 1957 paper.
Ibid. Inhibition of transport in the squid axon by ouabain had not been reported by 1958, and studies on squid were Skou’s link to the transport field.
Post (1959).
Post et al. (1960).
Ibid., p. 1801.
Aldridge (1962).
Post et al. (1960), pp. 1801, 1802.
Ibid. p. 1796.
Ibid.
Dunham and Glynn (1960), p. 62P.
Dunham and Glynn (1961).
Tosteson et al. (1960).
Hoffman (1961, 1962b ). The meeting abstract of 1960, by Hoffman and H. E. Ryan, was not published.
Skou (1961); Post and Albright (1961).
Bonting et al. (1961).
Bonting and Caravaggio (1963).
Glynn (1962); Whittam (1962a,b). Whittam had moved from Cambridge to Oxford.
Glynn (1962), p. 1813.
Post and Jolly (1957); Keynes and Swan (1959).
Sen and Post (1964). Others proposed that far fewer Na+ were transported for each ATP consumed, e.g., Caldwell et al. (1960a).
Skou (1960). It was submitted for publication in November 1959.
Charnock and Post (1963); a more complete account was published in Charnock et al. (1963).
The critical advance was including unlabeled ATP in the solution used to wash the precipitated enzyme after labeling with 32P-ATP.
Post et al. (1965).
Albers et al. (1963).
Fahn et al. (1964).
Glynn and Chappell (1964).
Glynn et al. (1965).
Hokin et al. (1965).
Nagano et al. (1965).
Glynn (1989b).
Quoted in Post (1989).
Perhaps that early misidentification contributed to Skou’s long disbelief in a phosphorylated enzyme participating in the ATPase reaction sequence (other evidence against E-P participating included an absence of stimulation by K+ at 0°).
Skou (1989), p. 436.
Post et al. (1960), p. 1796. On the other hand, not all those active in the field were then convinced of a stoichiometric link between Na+ outflux and K+ influx.
Deul and Mcllwain (1961), p. 246.
Schwartz et al. (1962), p. 626.
Tobias (1959), p. 306.
Altman (1959), p. 937.
For example, the increment in ATPase activity on adding Na+ and K+ was reported to be a mere 20% to 30% by Yoshida and Fujisawa (1962) and Emmelot and Bos (1962).
Hodgkin and Keynes (1955a, 1956). Skou (1989) acknowledged his ignorance of the latter paper at that time.
Skou (1960). Crab nerve ATPase required about 0.3 mM ouabain for half-maximal inhibition, several orders of magnitude more than needed for red blood cell ATPase. When an enzyme is inhibitable only by unusually high concentrations, there is a real likelihood that the effect is “non-specific,” i.e., that the reagent is then acting in a different fashion.
At a party after a friend’s thesis defense, Skou met Ejnar Lundsgaard, who was in Aarhus as an examiner; Skou mentioned to Lundsgaard his wish to prepare acetylcholinesterase and Lundsgaard arranged the visit with Nachmansohn, who was an old acquaintance.
For example, Kimura and DuBois (1947), in a paper on cardiotonic steroids and heart ATPase activity, failed to specify not only the monovalent cation content, but also the ATP concentration, the presence or absence of Mg, the reaction pH, and temperature.
Post, interview (1993).
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© 1997 American Physiological Society
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Robinson, J.D. (1997). Identifying the Na+/K+-ATPase. In: Moving Questions. People and Ideas Series. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-7600-9_9
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