Membrane-Associated Proteins and Peptides

Infrared spectroscopy is a useful technique for the determination of conformation and orientation of membrane-associated proteins and lipids. The technique is especially powerful for detecting conformational changes by recording spectral differences before and after perturbations in physiological solution. Polarized infrared measurements on oriented membrane samples have revealed valuable information on the orientation of chemical groupings and substructures within membrane molecules which is difficult to obtain by other methods. The application of infrared spectroscopy to the static and dynamic structure of proteins and peptides in lipid bilayers is reviewed with some emphasis on the importance of sample preparation. Limitations of the technique with regard to the absolute determination of secondary structure and orientation and new strategies for structural assignments are also discussed.

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Liposomes as Carriers of Membrane‐Associated Proteins and Peptides for Mass Spectrometric Analysis

Angewandte Chemie ◽

10.1002/ange.202103931 ◽

2021 ◽

Author(s):

Melissa Frick ◽

Christian Schwieger ◽

Carla Schmidt

Keyword(s):

Mass Spectrometric ◽

Mass Spectrometric Analysis ◽

Spectrometric Analysis ◽

Associated Proteins ◽

Proteins And Peptides

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Photoreceptor disk membrane substructures by means of the complementary freeze-fracture-replica methods

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100081644 ◽

1978 ◽

Vol 36 (2) ◽

pp. 156-157

Author(s):

A. Tonosaki ◽

M. Yamasaki ◽

H. Washioka ◽

J. Mizoguchi

Keyword(s):

Cell Membranes ◽

Freeze Fracture ◽

Purple Membrane ◽

Remarkable Case ◽

Single Face ◽

Disk Membrane ◽

Associated Proteins ◽

Photoreceptor Membranes

A vertebrate disk membrane is composed of 40 % lipids and 60 % proteins. Its fracture faces have been classed into the plasmic (PF) and exoplasmic faces (EF), complementary with each other, like those of most other types of cell membranes. The hypothesis assuming the PF particles as representing membrane-associated proteins has been challenged by serious questions if they in fact emerge from the crystalline formation or decoration effects during freezing and shadowing processes. This problem seems to be yet unanswered, despite the remarkable case of the purple membrane of Halobacterium, partly because most observations have been made on the replicas from a single face of specimen, and partly because, in the case of photoreceptor membranes, the conformation of a rhodopsin and its relatives remains yet uncertain. The former defect seems to be partially fulfilled with complementary replica methods.

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Characterization of microtubules by dark-field STEM and replication

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010008506x ◽

1991 ◽

Vol 49 ◽

pp. 152-153

Author(s):

S.B. Andrews ◽

R.D. Leapman ◽

P.E. Gallant ◽

T.S. Reese

Keyword(s):

Protein Interactions ◽

Low Dose ◽

Unit Length ◽

Dark Field ◽

Carbon Films ◽

Microtubule Associated Proteins ◽

Molecular Weights ◽

Freeze Dried ◽

Protein Motors ◽

Associated Proteins

As part of a study on protein interactions involved in microtubule (MT)-based transport, we used the VG HB501 field-emission STEM to obtain low-dose dark-field mass maps of isolated, taxol-stabilized MTs and correlated these micrographs with detailed stereo images from replicas of the same MTs. This approach promises to be useful for determining how protein motors interact with MTs. MTs prepared from bovine and squid brain tubulin were purified and free from microtubule-associated proteins (MAPs). These MTs (0.1-1 mg/ml tubulin) were adsorbed to 3-nm evaporated carbon films supported over Formvar nets on 600-m copper grids. Following adsorption, the grids were washed twice in buffer and then in either distilled water or in isotonic or hypotonic ammonium acetate, blotted, and plunge-frozen in ethane/propane cryogen (ca. -185 C). After cryotransfer into the STEM, specimens were freeze-dried and recooled to ca.-160 C for low-dose (<3000 e/nm2) dark-field mapping. The molecular weights per unit length of MT were determined relative to tobacco mosaic virus standards from elastic scattering intensities. Parallel grids were freeze-dried and rotary shadowed with Pt/C at 14°.

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Filaments and membranes in the mitotic apparatus

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010007638x ◽

1983 ◽

Vol 41 ◽

pp. 544-547

Author(s):

Kent McDonald

Keyword(s):

Intermediate Filaments ◽

Mitotic Spindle ◽

Mitotic Apparatus ◽

Light Microscope Level ◽

Microtubule Associated Proteins ◽

Recent Developments ◽

Associated Proteins ◽

Force Generator ◽

Spindle Function ◽

Antibody Technology

At the light microscope level the recent developments and interest in antibody technology have permitted the localization of certain non-microtubule proteins within the mitotic spindle, e.g., calmodulin, actin, intermediate filaments, protein kinases and various microtubule associated proteins. Also, the use of fluorescent probes like chlorotetracycline suggest the presence of membranes in the spindle. Localization of non-microtubule structures in the spindle at the EM level has been less rewarding. Some mitosis researchers, e.g., Rarer, have maintained that actin is involved in mitosis movements though the bulk of evidence argues against this interpretation. Others suggest that a microtrabecular network such as found in chromatophore granule movement might be a possible force generator but there is little evidence for or against this view. At the level of regulation of spindle function, Harris and more recently Hepler have argued for the importance of studying spindle membranes. Hepler also believes that membranes might play a structural or mechanical role in moving chromosomes.

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