Limits...
Membrane-directed molecular assembly of the neuronal SNARE complex.

Cho WJ, Lee JS, Zhang L, Ren G, Shin L, Manke CW, Potoff J, Kotaria N, Zhvania MG, Jena BP - J. Cell. Mol. Med. (2011)

Bottom Line: Using high-resolution electron microscopy, the electron density maps and 3D topography of the membrane-directed SNARE ring complex was determined at nanometre resolution.Furthermore, the mathematical prediction of the SNARE ring complex size with reasonable accuracy, and the possible mechanism of membrane-directed t-/v-SNARE ring complex assembly, was determined from the study.Therefore in the present study, using both lipososome-reconstituted recombinant t-/v-SNARE proteins, and native v-SNARE present in isolated SV membrane, the membrane-directed molecular assembly of the neuronal SNARE complex was determined for the first time and its size mathematically predicted.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Wayne State University School of Medicine, Detroit, MI 48201, USA.

Show MeSH

Related in: MedlinePlus

The possible establishment of a leak-proof SNARE ring complex channel is demonstrated (A). Size of the t-/v-SNARE ring complex is directly proportional to the size of the SNARE-associated vesicle (B). Different sizes of v-SNARE-associated vesicles, when interacting with t-SNARE-associated membrane (○), demonstrate the SNARE ring size to be directly proportional to the vesicle size. When a 50 nm diameter v-SNARE-reconstituted vesicle interacts with a t-SNARE-reconstituted membrane, a 11 nm diameter t-/v-SNARE ring complex is formed. Similarly, the present study demonstrates that when a 50 nm diameter v-SNARE-reconstituted vesicle, interacts with a 50 nm diameter t-SNARE-reconstituted vesicle, an 8 nm diameter t-/v-SNARE ring complex is established (♦). Analogous to the 11 nm diameter t-/v-SNARE ring complexes formed when 50 nm v-SNARE vesicles meet a t-SNARE-reconstituted planer membrane (B), approximately 11 nm diameter t-/v-SNARE ring complexes are formed when 50 nm diameter SVs meet a t-SNARE-reconstituted planer membrane (C, D).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3822491&req=5

fig05: The possible establishment of a leak-proof SNARE ring complex channel is demonstrated (A). Size of the t-/v-SNARE ring complex is directly proportional to the size of the SNARE-associated vesicle (B). Different sizes of v-SNARE-associated vesicles, when interacting with t-SNARE-associated membrane (○), demonstrate the SNARE ring size to be directly proportional to the vesicle size. When a 50 nm diameter v-SNARE-reconstituted vesicle interacts with a t-SNARE-reconstituted membrane, a 11 nm diameter t-/v-SNARE ring complex is formed. Similarly, the present study demonstrates that when a 50 nm diameter v-SNARE-reconstituted vesicle, interacts with a 50 nm diameter t-SNARE-reconstituted vesicle, an 8 nm diameter t-/v-SNARE ring complex is established (♦). Analogous to the 11 nm diameter t-/v-SNARE ring complexes formed when 50 nm v-SNARE vesicles meet a t-SNARE-reconstituted planer membrane (B), approximately 11 nm diameter t-/v-SNARE ring complexes are formed when 50 nm diameter SVs meet a t-SNARE-reconstituted planer membrane (C, D).

Mentions: To observe physiological SNARE complexes on mica for AFM studies, freshly cleaved mica disks were placed in a fluid chamber. Two hundred microlitres of imaging buffer solution containing 140 mM NaCl, 10 mM HEPES and 1 mM CaCl2, was placed at the centre of the cleaved mica disk. Prior to deposition on mica, t-SNARE reconstituted vesicles were gently layered on the mica surface, and incubated at room temperature for 60 min. prior to washing. Isolated SVs were added, incubated for 20 min., prior to three washes using the imaging buffer. During repeated imaging of the SVs attached to the t-SNARE-membrane supported by mica, some SVs are dislodged by the AFM cantilever tip, exposing the t-/v-SNARE ring complex (Fig. 5D).


Membrane-directed molecular assembly of the neuronal SNARE complex.

Cho WJ, Lee JS, Zhang L, Ren G, Shin L, Manke CW, Potoff J, Kotaria N, Zhvania MG, Jena BP - J. Cell. Mol. Med. (2011)

The possible establishment of a leak-proof SNARE ring complex channel is demonstrated (A). Size of the t-/v-SNARE ring complex is directly proportional to the size of the SNARE-associated vesicle (B). Different sizes of v-SNARE-associated vesicles, when interacting with t-SNARE-associated membrane (○), demonstrate the SNARE ring size to be directly proportional to the vesicle size. When a 50 nm diameter v-SNARE-reconstituted vesicle interacts with a t-SNARE-reconstituted membrane, a 11 nm diameter t-/v-SNARE ring complex is formed. Similarly, the present study demonstrates that when a 50 nm diameter v-SNARE-reconstituted vesicle, interacts with a 50 nm diameter t-SNARE-reconstituted vesicle, an 8 nm diameter t-/v-SNARE ring complex is established (♦). Analogous to the 11 nm diameter t-/v-SNARE ring complexes formed when 50 nm v-SNARE vesicles meet a t-SNARE-reconstituted planer membrane (B), approximately 11 nm diameter t-/v-SNARE ring complexes are formed when 50 nm diameter SVs meet a t-SNARE-reconstituted planer membrane (C, D).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3822491&req=5

fig05: The possible establishment of a leak-proof SNARE ring complex channel is demonstrated (A). Size of the t-/v-SNARE ring complex is directly proportional to the size of the SNARE-associated vesicle (B). Different sizes of v-SNARE-associated vesicles, when interacting with t-SNARE-associated membrane (○), demonstrate the SNARE ring size to be directly proportional to the vesicle size. When a 50 nm diameter v-SNARE-reconstituted vesicle interacts with a t-SNARE-reconstituted membrane, a 11 nm diameter t-/v-SNARE ring complex is formed. Similarly, the present study demonstrates that when a 50 nm diameter v-SNARE-reconstituted vesicle, interacts with a 50 nm diameter t-SNARE-reconstituted vesicle, an 8 nm diameter t-/v-SNARE ring complex is established (♦). Analogous to the 11 nm diameter t-/v-SNARE ring complexes formed when 50 nm v-SNARE vesicles meet a t-SNARE-reconstituted planer membrane (B), approximately 11 nm diameter t-/v-SNARE ring complexes are formed when 50 nm diameter SVs meet a t-SNARE-reconstituted planer membrane (C, D).
Mentions: To observe physiological SNARE complexes on mica for AFM studies, freshly cleaved mica disks were placed in a fluid chamber. Two hundred microlitres of imaging buffer solution containing 140 mM NaCl, 10 mM HEPES and 1 mM CaCl2, was placed at the centre of the cleaved mica disk. Prior to deposition on mica, t-SNARE reconstituted vesicles were gently layered on the mica surface, and incubated at room temperature for 60 min. prior to washing. Isolated SVs were added, incubated for 20 min., prior to three washes using the imaging buffer. During repeated imaging of the SVs attached to the t-SNARE-membrane supported by mica, some SVs are dislodged by the AFM cantilever tip, exposing the t-/v-SNARE ring complex (Fig. 5D).

Bottom Line: Using high-resolution electron microscopy, the electron density maps and 3D topography of the membrane-directed SNARE ring complex was determined at nanometre resolution.Furthermore, the mathematical prediction of the SNARE ring complex size with reasonable accuracy, and the possible mechanism of membrane-directed t-/v-SNARE ring complex assembly, was determined from the study.Therefore in the present study, using both lipososome-reconstituted recombinant t-/v-SNARE proteins, and native v-SNARE present in isolated SV membrane, the membrane-directed molecular assembly of the neuronal SNARE complex was determined for the first time and its size mathematically predicted.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Wayne State University School of Medicine, Detroit, MI 48201, USA.

Show MeSH
Related in: MedlinePlus