AN ALDOSTERONE REGULATED CHICKEN INTESTINE PROTEIN WITH HIGH-AFFINITYTO AMILORIDE

The pattern of chicken intestine amiloride-binding proteins was determ ined using the photoreactive amiloride analogue 2'-methoxy-5'-nitroben zamil (NMBA) and a polyclonal anti-amiloride antibody. At 10(-7)M, NMB A inhibits similar to 62% of the Na+ channel activity. At this concent ration the amiloride analogue labels a number of membrane proteins, an d in particular a 40-45 kDa polypeptide denoted ABP40. Incorporation o f NMBA into ABP40 could be prevented by a 100-fold excess of benzamil, but not by a 1000-fold excess of 5-(N-ethyl-N-isopropyl)-amiloride. L abeling of ABP40 was intense in membranes derived from salt-deprived c hickens and similar to 5-fold weaker in membranes from salt-repleted a nimals. Because of its small size, ABP40 is not likely to be an avian Na+ channel subunit, yet this amiloride-binding protein could be invol ved in the response to aldosterone. (C) 1997 Elsevier Science Inc.