Membrane asymmetry. A survey and critical appraisal of the methodology. I. Methods for assessing the asymmetric orientation and distribution of proteins.
This and the companion article are aimed at surveying the methods used for the study of membrane asymmetry. The techniques employed for the assessment of the asymmetric distribution and orientation of membrane proteins are reviewed in this article, whereas those pertaining to the unequal distribution of lipids are detailed in the companion paper. The use of immunological techniques and lectins, functions of proteins and their perturbations, chemical reagents, enzymatic isotopic labeling and enzymatic cleavage of membrane proteins and physical techniques are discussed and illustrated using recent examples of their application. Whenever appropriate, problems involving crypticity and non-availability or non-reactivity of functional sites, relevant chemical functions or protein fragments to appropriate ligands, reagents or modifying enzymes are envisaged and possible modification of the exposure of proteins during preparation of ghosts and other drawbacks are discussed, the use of different techniques and control experiments in conjunction is recommended for a more realistic assessment of the distribution and orientation of proteins.
D000818:Animals; D000936:Antigen-Antibody Complex; D005456:Fluorescent Dyes; D006241:Haptens; D007118:Immunoassay; D007123:Immunoelectrophoresis, Two-Dimensional / Q000379:methods; D007124:Immunoenzyme Techniques; D037102:Lectins; D008565:Membrane Proteins / Q000032:analysis* / Q000378:metabolism; D008566:Membranes / Q000648:ultrastructure*; D008854:Microscopy, Electron / Q000379:methods; D008856:Microscopy, Fluorescence
D000936:Antigen-Antibody Complex; D005456:Fluorescent Dyes; D006241:Haptens; D037102:Lectins; D008565:Membrane Proteins