Manijeh Mohraz, Ph.D.

Reconstitution of detergent-solubilized Na,K-ATPase and formation of two-dimensional crystals
Mohraz M
1999 Mar;125(1):76-85, Journal of structural biology
Very pure, detergent-solubilized Na,K-ATPase from dog or lamb kidneys has been successfully reconstituted at high protein-to-lipid weight ratios. Studies have been conducted to establish the orientation of the Na,K-ATPase molecules in the reconstituted membranes and to assess the functional activity and the conformational state of the reconstituted enzyme. Results indicate that reincorporation of the Na,K-ATPase molecules in the lipid bilayer is unidirectional and that the reconstituted enzyme retains its functional and structural integrity. Two-dimensional crystals have been induced in these preparations by vanadate ions. The arrays, with a dimeric structure in the unit cell, have a morphology similar to that of the crystals that had previously formed in the native membranes. Filtered images show that in projection, the molecule had an asymmetrical mass distribution, which at the resolution of 2.5 nm is identical to that of the earlier crystals. These sheets, although small, represent the first crystals of Na, K-ATPase to be formed by reconstitution. We expect that optimization of the reconstitution and crystallization parameters will lead to larger and better-ordered sheets, suitable for electron crystallography.
— id: 8513, year: 1999, vol: 125, page: 76, stat: Journal Article,

Reconstitution and crystallization of solubilized Na,K-ATPase
Mohraz, M
1999 JAN ;76(1):A451-A451, Biophysical journal
— id: 54000, year: 1999, vol: 76, page: A451, stat: Journal Article,

Production of recombinant Na1K-ATPase for structural studies
Mohraz, M; Morimoto, T
1999 NOV ;10(5):80A-80A, Molecular biology of the cell
— id: 53777, year: 1999, vol: 10, page: 80A, stat: Journal Article,

Reconstitution and crystallization of solubilized Na,K-ATPase
Mohraz, M
1997 NOV ;8(5):484-484, Molecular biology of the cell
— id: 53161, year: 1997, vol: 8, page: 484, stat: Journal Article,

Immunoelectron microscopy of epitopes on Na,K-ATPase catalytic subunit. Implications for the transmembrane organization of the C-terminal domain
Mohraz M; Arystarkhova E; Sweadner KJ
1994 Jan 28;269(4):2929-2936, Journal of biological chemistry
The transmembrane folding of the alpha subunit of Na,K-ATPase was studied by using immunoelectron microscopy to determine whether monoclonal antibodies with defined epitopes bind to the extracellular or cytoplasmic surface. In double labeling experiments, an antibody and a reference marker were bound to purified membrane-associated Na,K-ATPase and were visualized by employing colloidal gold particles of two different sizes. Wheat germ agglutinin and a previously characterized monoclonal antibody were used as control markers for the exoplasmic and cytoplasmic surfaces, respectively. Three antibodies, VG4, VG2, and IIC9, unambiguously bound to the extracellular surface. Previously IIC9 had been assigned to the cytoplasmic surface because, in immunofluorescence studies, it stained intact cells only when they were detergent-permeabilized. To investigate the basis for this contradiction, a third assay for sidedness was used: competition binding in solution to right-side-out renal medullary vesicles. IIC9 was found to bind to the extracellular surface of sealed vesicles, but only in certain experimental conditions. It was concluded that IIC9 has an epitope that is not always accessible and that in this instance, studies of binding to intact and detergent-treated cells had given misleading results. An extracellular disposition for all three antibodies is not compatible with existing folding models, and new models are presented
— id: 8430, year: 1994, vol: 269, page: 2929, stat: Journal Article,

MEMBRANE TOPOLOGY OF THE ALPHA SUBUNIT OF NA,K-ATPASE
MOHRAZ, M; ARYSTARKHOVA, E; SWEADNER, KJ
1993 FEB ;64(2):A331-A331, Biophysical journal
— id: 54341, year: 1993, vol: 64, page: A331, stat: Journal Article,

STRUCTURAL STUDY OF H,K-ATPASE BY ELECTRON-MICROSCOPY AND IMAGE-PROCESSING
Mohraz, M; Sathe, S; Smith, PR
1990 Feb;57(2):A357-A357, Biophysical journal
— id: 32111, year: 1990, vol: 57, page: A357, stat: Journal Article,

Three-dimensional structure of Na,K-ATPase and a model for the oligomeric form and the mechanism of the Na,K pump
Mohraz M; Smith PR
1988 ;268A:99-106, Progress in clinical & biological research
— id: 11284, year: 1988, vol: 268A, page: 99, stat: Journal Article,

The three-dimensional structure of the Na,K-ATPase from electron microscopy
Mohraz M; Simpson MV; Smith PR
1987 Jul;105(1):1-8, Journal of cell biology
The structure of Na,K-ATPase has been studied by electron microscopy and image reconstruction. A three-dimensional structure of this enzyme has been obtained to an overall resolution of 2.5 nm using data from specimens of negatively stained dimer sheets tilted through a range of angles +/- 60 degrees. The reconstruction shows a complex mass distribution consisting of ribbons of paired molecules extending approximately 6.0 nm from the cytoplasmic side of the membrane. The molecular envelope consists of a massive 'body' with 'lobe' and 'arm' structures projecting from it. The body has a columnar shape and is tilted with respect to the plane of the membrane. The region of interaction responsible for dimer formation is located between two bodies and is clearly visible in the reconstruction. It has been identified as a segment in the amino-terminal portion of the alpha subunit. The arms that interconnect the ribbons are located close to the membrane and are most probably formed by the beta subunits
— id: 8309, year: 1987, vol: 105, page: 1, stat: Journal Article,

3-D STRUCTURE OF NA,K-ATPASE BY ELECTRON-MICROSCOPY AND IMAGE- RECONSTRUCTION
Mohraz, M; Smith, PR
1987 Feb;51(2):A174-A174, Biophysical journal
— id: 31413, year: 1987, vol: 51, page: A174, stat: Journal Article,

The structure of the adenovirus capsid. III. Hexon packing determined from electron micrographs of capsid fragments
van Oostrum, J; Smith, P R; Mohraz, M; Burnett, R M
1987 Nov 5;198(1):73-89, Journal of molecular biology
The orientation and relative positions of all 240 hexons in the icosahedral outer capsid of adenovirus have been determined. Two types of capsid fragments, obtained after selective disruption of the virion, were analyzed using electron microscopy and image-processing techniques. Planar inverted groups-of-nine, arising from the central region of the capsid facet, were minimally stained to reveal the morphology of restricted regions of their component hexons. Images shown to be related by correspondence analysis were averaged and features of the individual hexon molecule, known from an X-ray crystallographic investigation, were used in their interpretation. The study confirms earlier observations that the hexons in the group-of-nine are distributed on a p3 net, shows that the hexons form a close-packed array using the pseudo-hexagonal shape of the hexon base, and provides their relative positions. Twenty interlocking groups-of-nine account for 180 of the 240 hexons present in the viral capsid. The orientation of the remaining 60 peripentonal hexons was obtained from a rotationally averaged image of a quarter-capsid, a novel viral fragment comprising five complete facets. Each peripentonal hexon forms planar asymmetric interactions with two neighbors in an adjacent group-of-nine so that it lies on an extension of the p3 net. The complete facet thus consists of 12 hexons arranged on a planar p3 net, with a shape that permits interlocking of hexons at the capsid edge. The relative positions of the hexons have been determined to within 5 A using the molecular model, and indicate that the pseudo-hexagonal basal regions are close-packed in a manner that maximizes the hexon-hexon contacts. The results confirm the model proposed earlier for the arrangement of hexons within the adenovirus capsid (Burnett, 1985), and show the power of the inter-disciplinary approach
— id: 150636, year: 1987, vol: 198, page: 73, stat: Journal Article,

THE VERTEX MORPHOLOGY OF THE ADENOVIRUS CAPSID
Vanoostrum, J; Smith, PR; Mohraz, M; Burnett, RM
1987 May 27;494(11):423-426, Annals of the New York Academy of Sciences
— id: 31309, year: 1987, vol: 494, page: 423, stat: Journal Article,

Structural studies of Na,K-ATPase
Mohraz, M; Yee, M; Smith, P R
1986 ;483:131-139, Annals of the New York Academy of Sciences
— id: 150635, year: 1986, vol: 483, page: 131, stat: Journal Article,

Interpretation of electron micrographs of adenovirus hexon arrays using a crystallographic molecular model
van Oostrum, J; Smith, P R; Mohraz, M; Burnett, R M
1986 Jul-Sep;96(1-3):77-90, Journal of ultrastructure & molecular structure research
Two types of two-dimensional arrays of purified adenovirus type 2 hexon have been obtained and analyzed by Fourier filtration of their electron micrographs. One array contained continuously close-packed hexons, distributed on a hexagonal p3 lattice, with a unit cell dimension of 94 +/- 2 A. The other array contained close-packed hexons with a regular absence, so that rings of six hexons related by sixfold symmetry formed a p6 unit cell. The cell dimension of the hexagonal array was 153 +/- 3 A, with neighboring hexons separated by 88 +/- 2 A. Smaller p6 arrays were also formed by hexons freed from complete virions on the microscope grid by treatment with distilled water. A molecular model of hexon, known from the X-ray crystallographic structure, was used to interpret Fourier-filtered images of the arrays, and to determine the relative orientations of the hexon molecules. The hexon-hexon interaction in the p3 array is that found in the virion facet, whereas that in the p6 array is a planar form of the interaction between peripentonal hexons around the vertex
— id: 150637, year: 1986, vol: 96, page: 77, stat: Journal Article,

Novel crystalline sheets of Na,K-ATPase induced by phospholipase A2
Mohraz M; Yee M; Smith PR
1985 Oct-Nov;93(1-2):17-26, Journal of ultrastructure research
Treatment of purified preparations of Na,K-ATPase by phospholipase A2 has led to the formation of two-dimensional crystals of the protein. Control tests with another phospholipase and two detergents have shown that crystallization occurs as the result of hydrolysis and/or solubilization of the phospholipids in the enzyme vesicles. Experimentation with various buffer systems has indicated that reduction in the amount of phospholipids alone is sufficient for inducing the formation of crystalline sheets. Inclusion of crystal inducing ions in the buffer facilitates the crystallization process, resulting in more extensive arrays. The new crystalline sheets are exclusively dimeric with average unit cell dimensions: a = 15.8 +/- 0.4 nm, b = 4.9 +/- 0.2 nm, and gamma = 64 +/- 3 degrees. Examination of the micrographs shows that the initial intermolecular interaction leading to the formation of sheets is between the alpha subunits. Results from this study suggest that removal and/or modification of phospholipids by phospholipases could prove successful in crystallizing those membrane proteins in which excess lipid is the main barrier to the formation of two-dimensional arrays
— id: 57472, year: 1985, vol: 93, page: 17, stat: Journal Article,

PROGRESS IN THE 3-D STRUCTURE DETERMINATION OF NA-K-ATPASE BY ELECTRON-MICROSCOPY AND IMAGE-PROCESSING
Mohraz, M; Smith, PR
1985 ;47(2):A173-A173, Biophysical journal
— id: 30789, year: 1985, vol: 47, page: A173, stat: Journal Article,

Structure of (Na+,K+)-ATPase as revealed by electron microscopy and image processing
Mohraz M; Smith PR
1984 May;98(5):1836-1841, Journal of cell biology
(Na+,K+)-ATPase was studied by electron microscopy and image processing of negatively stained and freeze-dried and shadowed crystalline sheets induced by a number of inorganic salts. Extensive experiments have identified new conditions for optimum crystal formation. Two crystal forms have been observed, one with a monomer and the other with a dimer, in the unit cell. Both show the same structure for the enzyme monomer. The enzyme can also be crystallized after partial proteolysis of its alpha subunit by trypsin. The proteolysed enzyme crystallizes under the same conditions as the whole enzyme. Comparison of the mass distributions in the images of the intact and proteolysed enzyme has allowed the tentative identification of the location of the alpha subunit within the monomer. The relationship between the structure of the crystallized enzyme and that of the enzyme in its native form is discussed, as is its apparent close structural relationship to the calcium-ATPase
— id: 57473, year: 1984, vol: 98, page: 1836, stat: Journal Article,

THE STRUCTURE OF (NA+,K+)-ATPASE AS REVEALED BY ELECTRON-MICROSCOPY
MOHRAZ, M; RINDER, CA; SIMPSON, MV; SMITH, PR
1984 ;435(DEC):561-563, Annals of the New York Academy of Sciences
— id: 41242, year: 1984, vol: 435, page: 561, stat: Journal Article,

THE STRUCTURE OF (NA+,K+)-ATPASE AS REVEALED BY ELECTRON-MICROSCOPY
MOHRAZ, M; RINDER, CA; SIMPSON, MV; SMITH, PR
1983 ;97(5):A116-A116, Journal of cell biology
— id: 40606, year: 1983, vol: 97, page: A116, stat: Journal Article,