Direct visualization of phosphorylase-phosphorylase kinase complexes by scanning tunneling and atomic force microscopy

Edstrom, Ronald D.; Meinke, Marilyn H.; Yang, Xu; Yang, Ruiguo; Elings, Virgil B.; Evans, D. F.

doi:10.1016/s0006-3495(90)82489-9

Cited by 72 publications

(49 citation statements)

References 23 publications

(15 reference statements)

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“…Concentrations of ligands: 100 mM K ϩ , 5 mM IDP. smFRET measurement implicated that two N termini on homodimeric subunits are relatively distant from each other compared to a theoretical diameter of 77.8 Å (12,29). In contrast to C termini, N termini are possibly remote from the interface of homodimeric CtH ϩ -PPase.…”

Section: Table 1 Fret Efficiencies (E*) Of Mutant Cthmentioning

confidence: 97%

Distance Variations between Active Sites of H+-Pyrophosphatase Determined by Fluorescence Resonance Energy Transfer

Huang

Liu

Chen

et al. 2010

Journal of Biological Chemistry

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Section: Table 1 Fret Efficiencies (E*) Of Mutant Cthmentioning

confidence: 97%

Distance Variations between Active Sites of H+-Pyrophosphatase Determined by Fluorescence Resonance Energy Transfer

Huang

Liu

Chen

et al. 2010

Journal of Biological Chemistry

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“…where M 0 is the molecular mass of the protein, N 0 is Avogadro's number, V 1 and V 2 are the partial specific volumes of protein and water (0.74 and 1 cm 3 /g, respectively), and d is the extent of protein hydration (0.4 mol water/mol protein) (13).…”

Section: Formation Of Supported Lipidmentioning

confidence: 99%

Placental Alkaline Phosphatase Is Efficiently Targeted to Rafts in Supported Lipid Bilayers

Saslowsky¹,

Lawrence²,

Ren³

et al. 2002

Journal of Biological Chemistry

160

161

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Evidence is growing that biological membranes contain lipid microdomains or "rafts" that may be involved in processes such as cellular signaling and protein trafficking. In this study, we have used atomic force microscopy to examine the behavior of rafts in supported lipid bilayers. We show that bilayers composed of equimolar dioleoylphosphatidylcholine and sphingomyelin spontaneously form rafts, which are detectable as raised features. A comparison of the extents of protrusion of the rafts in monolayers and bilayers indicates that the rafts in the two leaflets of the bilayer coincide. The rafts were observed both in the absence and presence of cholesterol (33 mol %). Cholesterol reduced raft protrusion presumably by increasing the thickness of the non-raft bilayer. PLAP (glycosylphosphatidylinositol-anchored protein placental alkaline phosphatase) was purified and shown to exist as a dimer. Following its incorporation into supported lipid bilayers, PLAP was found to be targeted efficiently to rafts, both in the absence and presence of cholesterol. We suggest that atomic force microscopy provides a powerful tool for the study of raft structure and properties.

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“…where M 0 is the molecular mass of the protein, N 0 is Avogadro's number, V 1 and V 2 are the partial speci®c volumes of protein and water (0.74 cm 3 g 71 and 1 cm 3 g 71 , respectively), and d is the extent of protein hydration (0.4 mol water/mol protein) (Edstrom et al, 1990).…”

Section: Molecular Volume Calculationmentioning

confidence: 99%

Direct visualization of ligand‐protein interactions using atomic force microscopy

Neish

Martin

Henderson

et al. 2002

British J Pharmacology

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1 Streptavidin is a 60-kDa tetramer which binds four molecules of biotin with extremely high anity (K A *10 14 M 71). We have used atomic force microscopy (AFM) to visualize this ligandprotein interaction directly. 2 Biotin was tagged with a short (152-basepair; 50-nm) DNA rod and incubated with streptavidin. The resulting complexes were then imaged by AFM. The molecular volume of streptavidin calculated from the dimensions of the protein particles (105+3 nm 3 ) was in close agreement with the value calculated from its molecular mass (114 nm 3 ). Biotinylation increased the apparent size of streptavidin (to 133+2 nm 3 ), concomitant with an increase in the thermal stability of the tetramer. 3 Images of streptavidin with one to four molecules of DNA-biotin bound were obtained. When two ligands were bound, the angle between the DNA rods was either acute or obtuse, as expected from the relative orientations of the biotin binding sites. The ratio of acute : obtuse angles (1 : 3) was lower than the expected value (1 : 2), indicating a degree of steric hindrance in the binding of the DNA-biotin. The slight under-representation of higher occupancy states supported this idea. 4 Streptavidin with a single molecule of DNA-biotin bound was used to tag biotinylated b-galactosidase, a model multimeric enzyme. 5 The ability to image directly the binding of a ligand to its protein target by AFM provides useful information about the nature of the interaction, and about the eect of complex formation on the structure of the protein. Furthermore, the use of DNA-biotin/streptavidin tags could potentially shed light on the architecture of multi-subunit proteins.

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Direct visualization of phosphorylase-phosphorylase kinase complexes by scanning tunneling and atomic force microscopy

Cited by 72 publications

References 23 publications

Distance Variations between Active Sites of H+-Pyrophosphatase Determined by Fluorescence Resonance Energy Transfer

Distance Variations between Active Sites of H+-Pyrophosphatase Determined by Fluorescence Resonance Energy Transfer

Placental Alkaline Phosphatase Is Efficiently Targeted to Rafts in Supported Lipid Bilayers

Direct visualization of ligand‐protein interactions using atomic force microscopy

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