Structural organization, ion transport, and energy transduction of P-type ATPases

Endoplasmic Reticulum Quality Control of Oligomeric Membrane Proteins: Topogenic Determinants Involved in the Degradation of the Unassembled Na,K-ATPase α Subunit and in Its Stabilization by β Subunit Assembly

“…ϩ, ␣-proteins degraded after a 48-h chase; Ϫ, ␣-proteins not degraded after a 48-h chase; M, membrane segment; nd, not determined. At the bottom, a putative, linear model of the ␣ subunit of Xenopus Na,K-ATPase is shown, which indicates the amino acids that define the putative transmembrane segments according to a 10-transmembrane segment model (Moller et al, 1996). points in the ␣ cDNA cloned into the pSD5 vector by using the PCR method (Nelson and Long, 1989).…”

“…To identify the critical amino acids involved, we tested the degradation of truncated M1-5 ␣-proteins of different length. According to a recently proposed topology model (Moller et al, 1996;Béguin et al, 1998), M5 of the ␣ subunit of Na,KATPase starts at Ser-777, ends at Ile-800, and is connected to M6 by a three-amino-acid-long extracellular loop containing two proline residues (see Table 1). M1-5 ␣-proteins ending at Ile-786 ( Figure 2A, lanes 1 and 2) or at Ala-798 (lanes 3 and 4) were stable during a 48-h chase similar to a M1-4 Q698 ␣-protein ( Figure 1A, lanes 7 and 8), whereas an M1-5 ␣-protein ending at Leu-804 was degraded ( Figure 2A, lanes 5 and 6) similar to the M1-5 G815 ␣-protein ( Figure 1A, lanes 9 and 10).…”

“…Our results indeed indicate that certain ␣ domains, which in the mature, ␤-assembled ␣ subunit are located on the extracytoplasmic side, are transiently exposed to the cytoplasmic side during synthesis of the ␣ protein because of inefficient membrane insertion of C-terminal membrane domains and potentially become targets for cytosolic, proteasomal attack (Figure 8). One of the candidate domains that could be implicated in such a degradation process comprises M5 and the M5/M6 extracytoplasmic loop, which contains of a Pro-Leu-Pro motif highly conserved in Na,K-and H,K-ATPases (Moller et al, 1996). According to mutational analysis, the two proline residues strongly impede membrane insertion of M5 (Bé-guin et al, 1998) and at the same time may be involved in the degradation of ␣ proteins from the cytosolic side (Figure 2) most likely by the proteasomal system ( Figure 5).…”

“…Finally, the example of the Na,K-ATPase ␣ subunit permits some speculation on the necessity of partner subunit assembly in the maturation of oligomeric transport proteins. In the case of P-type ATPases, transport function is achieved through conformational changes that involve the C-terminal part of the ␣ subunit (for review, see Moller et al, 1996). In contrast to other P-type ATPases, which do not need assembly with ␤ subunits for maturation, in Na,K-and H,KATPase ␣ subunits, the specific sequence information in the C-terminal membrane domains, which provide the conformational flexibility, may be so stringent that it requires, during evolution, the association of a helper protein to permit correct membrane insertion.…”

Mass spectrometry in aging research

“…SERCA plays a major role in muscle relaxation through the ATP-dependent transport of cytosolic calcium into the lumen of the sarcoplasmic reticulum (Møller, Juul, & le Maire, 1996;MacLennan, Rice, & Green, 1997). In general, calcium transport into the SR or ER constitutes an important part in the maintenance of intracellular calcium homeostasis together with calcium transport across the plasma membrane and into the mitochondria (Trump & Berezesky, 1995).…”