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Localization of intestinal sucrase-isomaltase complex on the microvillous membrane by electron microscopy using
Abstract:
Microvillous vesicles isolated from rabbit small intestine showed a trilaminar membrane with a rather smooth surface, which was apparently not affected by papain solubilizing sucrase-isomaltase complex or by trypsin unable to solubilize it. When microvilous vesicles or trysinized ones were incubated with immunoglobulin G against the sucrase-isomaltase complex or monovalent fragments therefrom, an apparently continuous electron-opaque layer approximately 180 A in width appeared around the external surface of vesicles. Such a layer was not formed on papainized vesicles. Microvillous vesicles and trypsinized ones negatively stained with phosphotungate showed a great number of particles protruding approximately 150 A from the membrane surface, but papainized vesicles did not. The particles existed close to one another and appeared to form a particulate layer 150 A in width on the surface. The antibodies, whether they were divalent or monovalent, increased the width of the layer to approximately 200 A and obscured the fine particulate structure of intact and trypsinized vesicles. Papainized vesicles retained their smooth surface upon interaction with antibodies. These results, together with those with the Triton-solubilized sucrase- isomaltase complex (Nishi and Takesue, 1978), J. Ultra-struct. Res., 62:1- 12), indicate not only that sucrase-isomaltase complexes are located close to one another on the membrane, but also that they or at least their protein portions protrude approximately 150 A from the surface of the trilaminar membrane.
Insights
Rabbit small intestine microvilli possess sucrase-isomaltase complexes that protrude from the membrane surface. Antibody binding to these complexes increases their apparent layer width, confirming their surface localization.
Area of Science:
- Biochemistry
- Cell Biology
- Gastroenterology
Background:
- The small intestine's microvilli are crucial for nutrient absorption.
- Sucrase-isomaltase complex is a key enzyme embedded in the microvillus membrane.
Purpose of the Study:
- To investigate the structural localization and surface protrusion of sucrase-isomaltase complexes on rabbit small intestine microvilli.
- To determine the effect of enzymatic treatment and antibody binding on the microvillus surface structure.
Main Methods:
- Isolation of microvillous vesicles from rabbit small intestine.
- Enzymatic treatment with papain and trypsin.
- Incubation with antibodies (immunoglobulin G) against the sucrase-isomaltase complex.
- Negative staining with phosphotungstate and electron microscopy.
Main Results:
- Intact and trypsinized vesicles showed a particulate surface layer approximately 150 A wide, attributed to sucrase-isomaltase complexes.
- Papain treatment removed this layer, indicating the complexes are papain-solubilizable.
- Antibody binding increased the apparent layer width to ~200 A and obscured particulate structure.
- Papainized vesicles remained smooth after antibody incubation.
Conclusions:
- Sucrase-isomaltase complexes are located on the microvillus membrane surface and protrude approximately 150 A.
- These complexes are closely packed and their protein portions are exposed.
- Enzymatic digestion and antibody interactions provide insights into the structural organization of microvillar enzymes.