title: Distinctive features of catalytic and transport mechanisms in mammalian sarco-endoplasmic reticulum Ca2+ ATPase (SERCA) and Cu+ (ATP7A/B) ATPases.
abstract: Ca(2+) (sarco-endoplasmic reticulum Ca(2+) ATPase (SERCA)) and Cu(+) (ATP7A/B) ATPases utilize ATP through formation of a phosphoenzyme intermediate (E-P) whereby phosphorylation potential affects affinity and orientation of bound cation. SERCA E-P formation is rate-limited by enzyme activation by Ca(2+), demonstrated by the addition of ATP and Ca(2+) to SERCA deprived of Ca(2+) (E2) as compared with ATP to Ca(2+)-activated enzyme (E1·2Ca(2+)). Activation by Ca(2+) is slower at low pH (2H(+)·E2 to E1·2Ca(2+)) and little sensitive to temperature-dependent activation energy. On the other hand, subsequent (forward or reverse) phosphoenzyme processing is sensitive to activation energy, which relieves conformational constraints limiting Ca(2+) translocation. A "H(+)-gated pathway," demonstrated by experiments on pH variations, charge transfer, and Glu-309 mutation allows luminal Ca(2+) release by H(+)/Ca(2+) exchange. As compared with SERCA, initial utilization of ATP by ATP7A/B is much slower and highly sensitive to temperature-dependent activation energy, suggesting conformational constraints of the headpiece domains. Contrary to SERCA, ATP7B phosphoenzyme cleavage shows much lower temperature dependence than EP formation. ATP-dependent charge transfer in ATP7A and -B is observed, with no variation of net charge upon pH changes and no evidence of Cu(+)/H(+) exchange. As opposed to SERCA after Ca(2+) chelation, ATP7A/B does not undergo reverse phosphorylation with P(i) after copper chelation unless a large N-metal binding extension segment is deleted. This is attributed to the inactivating interaction of the copper-deprived N-metal binding extension with the headpiece domains. We conclude that in addition to common (P-type) phosphoenzyme intermediate formation, SERCA and ATP7A/B possess distinctive features of catalytic and transport mechanisms.
authors.affiliation: California Pacific Medical Center Research Institute, San Francisco, California 94107, USA.
authors.forename: David
authors.initials: D
authors.lastname: Lewis
authors.forename: Rajendra
authors.initials: R
authors.lastname: Pilankatta
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Inesi
authors.forename: Gianluca
authors.initials: G
authors.lastname: Bartolommei
authors.forename: Maria Rosa
authors.initials: MR
authors.lastname: Moncelli
authors.forename: Francesco
authors.initials: F
authors.lastname: Tadini-Buoninsegni
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D002384:Catalysis; D027682:Cation Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D000073840:Copper-Transporting ATPases; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D017136:Ion Transport / Q000502:physiology; D017434:Protein Structure, Tertiary; D011817:Rabbits; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D027682:Cation Transport Proteins; D000251:Adenosine Triphosphatases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; C000612464:ATP7B protein, human; D000073840:Copper-Transporting ATPases; D002118:Calcium