Two Disease-Causing SNAP-25B Mutations Selectively Impair SNARE C-terminal Assembly. Issue 4 (16th February 2018)

• Record Type: Journal Article
• Authors: Rebane, Aleksander A.; Wang, Bigeng; Ma, Lu; Qu, Hong; Coleman, Jeff; Krishnakumar, Shyam; Rothman, James E.; Zhang, Yongli
• Is Part Of: Journal of molecular biology. Volume 430:Issue 4(2018)
• Page Start: 479
• Page End: 490
• Publication Date: 2018-02-16
• Subjects: SNARE soluble N-ethylmaleimide-sensitive factor attachment receptors -- VAMP2 vesicle-associated membrane protein 2 -- SNAP-25 synaptosomal-associated protein of molecular weight 25 kDa -- v-SNARE vesicle-associated SNARE -- t-SNARE target membrane-associated SNARE -- NTD N-terminal domain of the SNARE complex -- CTD C-terminal domain of the SNARE complex -- LD linker domain of the SNARE complex -- FEC force–extension curve -- HMM hidden Markov modeling; optical tweezers -- SNARE assembly -- membrane fusion -- protein folding -- neuropathy; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2017.10.012
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: Synaptic exocytosis relies on assembly of three soluble N -ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins into a parallel four-helix bundle to drive membrane fusion. SNARE assembly occurs by stepwise zippering of the vesicle-associated SNARE (v-SNARE) onto a binary SNARE complex on the target plasma membrane (t-SNARE). Zippering begins with slow N-terminal association followed by rapid C-terminal zippering, which serves as a power stroke to drive membrane fusion. SNARE mutations have been associated with numerous diseases, especially neurological disorders. It remains unclear how these mutations affect SNARE zippering, partly due to difficulties to quantify the energetics and kinetics of SNARE assembly. Here, we used single-molecule optical tweezers to measure the assembly energy and kinetics of SNARE complexes containing single mutations I67T/N in neuronal SNARE synaptosomal-associated protein of 25 kDa (SNAP-25B), which disrupt neurotransmitter release and have been implicated in neurological disorders. We found that both mutations significantly reduced the energy of C-terminal zippering by ~ 10 k B T, but did not affect N-terminal assembly. In addition, we observed that both mutations lead to unfolding of the C-terminal region in the t-SNARE complex. Our findings suggest that both SNAP-25B mutations impair synaptic exocytosis by destabilizing SNARE assembly, rather than stabilizing SNARE assembly as previously proposed. Therefore, our measurements provide insights into the molecular mechanism of the disease caused by SNARE mutations. Graphical Abstract: Highlights: The mechanism by which two SNAP-25B mutations cause disease is unclear. The mutations greatly weaken SNARE C-terminal zippering. The mutations do not affect SNARE N-terminal assembly. The mutations impair t-SNARE folding. The mutations impair SNARE assembly and thus lead to impaired neurotransmission.