Monolayer freeze-fracture and scanning tunneling microscopy.
This article reviews research on planar monolayer methods, application of the methods to analyses of transmembrane signaling, and the combination of these methods with scanning tunneling microscopy (STM). Past research has involved the development of monolayer freeze-fracture methods. These include monolayer freeze-fracture autoradiography (MONOFARG), an electron microscopic cytochemical method to analyze in-plane distributions of radioisotopes, and double-labeled membrane splitting (DBLAMS) and single-membrane monolayer splitting (SMMS), methods to analyze transmembrane distributions of native and radiolabeled proteins and lipids. Present research has focussed on using these methods to investigate mechanisms of transmembrane signaling mediated by protein kinase C (PKC), including the transbilayer distribution of the tumor promoter TPA, a lipophilic activator of PKC, and the transbilayer distribution of peripheral membrane proteins phosphorylated by PKC. Future work will involve the combination of planar sample preparation with STM. The principles and applications of biological STM are briefly reviewed and a low-resolution STM image of a planar purple-membrane monolayer is included. The combination of planar methods and STM can provide the chemical information lacking in STM images enabling microscopists to investigate biochemical phenomena at nanometer resolution.
Fisher
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Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0130.
D001345:Autoradiography; D005614:Freeze Fracturing / Q000379:methods*; D006801:Humans; D008566:Membranes / Q000648:ultrastructure; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D011493:Protein Kinase C / Q000378:metabolism; D015398:Signal Transduction; D013048:Specimen Handling
D011493:Protein Kinase C