Three-Dimensional Structure of the Porcine Gastric H,K-ATPase from Negatively Stained Crystals

Xian, Yijing; Hebert, Hans

doi:10.1006/jsbi.1997.3847

Cited by 15 publications

(9 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In any case, the principles developed in this work are relevant to other membrane proteins and, in particular, to the other members of the P-type ion pump family that are related in structure and function to Ca-ATPase. In this context, vanadate and phospholipase A2-induced 2D crystals of Na,K-ATPase, H,K-ATPase, and Kdp-ATPase have already been obtained in native membranes (Hebert et al, 1985;Mohraz et al, 1985;Iwane et al, 1996;Xian and Hebert, 1997), but are poorly ordered. Future structural analysis of these P-type ATPases will require crystallization by reconstitution from detergentsolubilized protein.…”

Section: Resultsmentioning

confidence: 99%

Two-Dimensional Crystallization of Ca-ATPase by Detergent Removal

Lacapère

Stokes

Olofsson

et al. 1998

Biophysical Journal

View full text Add to dashboard Cite

By using Bio-Beads as a detergent-removing agent, it has been possible to produce detergent-depleted two-dimensional crystals of purified Ca-ATPase. The crystallinity and morphology of these different crystals were analyzed by electron microscopy under different experimental conditions. A lipid-to-protein ratio below 0.4 w/w was required for crystal formation. The rate of detergent removal critically affected crystal morphology, and large multilamellar crystalline sheets or wide unilamellar tubes were generated upon slow or fast detergent removal, respectively. Electron crystallographic analysis indicated unit cell parameters of a = 159 A, b = 54 A, and gamma = 90 degrees for both types of crystals, and projection maps at 15-A resolution were consistent with Ca-ATPase molecules alternately facing the two sides of the membrane. Crystal formation was also affected by the protein conformation. Indeed, tubular and multilamellar crystals both required the presence of Ca2+; the presence of ADP gave rise to another type of packing within the unit cell (a = 86 A, b = 77 A, and gamma = 90 degrees), while maintaining a bipolar orientation of the molecules within the bilayer. All of the results are discussed in terms of nucleation and crystal growth, and a model of crystallogenesis is proposed that may be generally true for asymmetrical proteins with a large hydrophilic cytoplasmic domain.

show abstract

Section: Resultsmentioning

confidence: 99%

Two-Dimensional Crystallization of Ca-ATPase by Detergent Removal

Lacapère

Stokes

Olofsson

et al. 1998

Biophysical Journal

View full text Add to dashboard Cite

show abstract

“…To investigate the conformational changes in P-type ATPases in the context of a lipid bilayer, we attempted to reconstitute H + ,K + -ATPase in different states of the reaction cycle into 2D crystals. It has previously been reported that vanadate induces the formation of 2D crystals of the H + ,K + -ATPase in the native, gastric membrane (Rabon et al, 1986;Xian and Hebert, 1997), but larger and betterordered 2D crystals would be desirable for structural analyses at higher resolution. Additionally these crystals had not been analyzed by cryo-electron microscopy (cryo-EM).…”

Section: Introductionmentioning

confidence: 98%

Structural analysis of 2D crystals of gastric H+,K+-ATPase in different states of the transport cycle

Nishizawa

Abe

Tani

et al. 2008

Journal of Structural Biology

View full text Add to dashboard Cite

“…These membranes have been used for numerous molecular biological, biochemical and biophysical studies and early enzyme preparations proved that this family of ion pumps was particularly amenable to techniques of structural biology (Skriver, Maunsbach & Jorgensen, 1981;Dux & Martonosi, 1983;Rabon et al, 1986). While initial insights into the structural mechanism of transport were obtained by electron microscopy (Xian & Hebert, 1997;Auer, Scarborough & Kuhlbrandt, 1998;Ogawa et al, 1998;Zhang et al, 1998), recent x-ray crystallographic studies have made the final step to atomic resolution.…”

Section: Introductionmentioning

confidence: 99%