Liming Yan

Magnetoelectric coupling in the paramagnetic state of a metal-organic framework

Although the magnetoelectric effects - the mutual control of electric polarization by magnetic fields and magnetism by electric fields, have been intensively studied in a large number of inorganic compounds and heterostructures, they have been rarely observed in organic materials. Here we demonstrate magnetoelectric coupling in a metal-organic framework [(CH3)2NH2]Mn(HCOO)3 which exhibits an order-disorder type of ferroelectricity below 185 K. The magnetic susceptibility starts to deviate from the Curie-Weiss law at the paraelectric-ferroelectric transition temperature, suggesting an enhancement of short-range magnetic correlation in the ferroelectric state. Electron spin resonance study further confirms that the magnetic state indeed changes following the ferroelectric phase transition. Inversely, the ferroelectric polarization can be improved by applying high magnetic fields. We interpret the magnetoelectric coupling in the paramagnetic state in the metal-organic framework as a consequence of the magnetoelastic effect that modifies both the superexchange interaction and the hydrogen bonding.

DWARF14 is a non-canonical hormone receptor for strigolactone

Classical hormone receptors reversibly and non-covalently bind active hormone molecules, which are generated by biosynthetic enzymes, to trigger signal transduction. The α/β hydrolase DWARF14 (D14), which hydrolyses the plant branching hormone strigolactone and interacts with the F-box protein D3/MAX2, is probably involved in strigolactone detection. However, the active form of strigolactone has yet to be identified and it is unclear which protein directly binds the active form of strigolactone, and in which manner, to act as the genuine strigolactone receptor. Here we report the crystal structure of the strigolactone-induced AtD14–D3–ASK1 complex, reveal that Arabidopsis thaliana (At)D14 undergoes an open-to-closed state transition to trigger strigolactone signalling, and demonstrate that strigolactone is hydrolysed into a covalently linked intermediate molecule (CLIM) to initiate a conformational change of AtD14 to facilitate interaction with D3. Notably, analyses of a highly branched Arabidopsis mutant d14-5 show that the AtD14(G158E) mutant maintains enzyme activity to hydrolyse strigolactone, but fails to efficiently interact with D3/MAX2 and loses the ability to act as a receptor that triggers strigolactone signalling in planta. These findings uncover a mechanism underlying the allosteric activation of AtD14 by strigolactone hydrolysis into CLIM, and define AtD14 as a non-canonical hormone receptor with dual functions to generate and sense the active form of strigolactone.

Structural basis for the impact of phosphorylation on the activation of plant receptor-like kinase BAK1.

Structural basis for the impact of phosphorylation on the activation of plant receptor-like kinase BAK1

The Crystal Structure of Porcine Reproductive and Respiratory Syndrome Virus Nonstructural Protein Nsp1β Reveals a Novel Metal-Dependent Nuclease

ABSTRACT Porcine reproductive and respiratory syndrome virus (PRRSV), a member of the Arteriviridae family of Nidovirales, is the causative agent of porcine reproductive and respiratory syndrome, which results in enormous economic losses in the swine industry. As the second protein encoded by the PRRSV genome, nsp1β cleaves itself from the downstream nsp2 protein via a C-terminal papain-like cysteine protease (PCP) domain. Although nsp1β is known to be involved in virulence, its precise role in the process of viral infection remains unclear. In this work, we describe the homodimeric crystal structure of PRRSV nsp1β in its natural, self-processed form. We show that the architecture of its N-terminal domain (NTD) adopts a fold closely resembling that of several known nucleases and has intrinsic nuclease activity that is strongly activated by manganese ions in vitro. Key features, however, distinguish nsp1β from characterized nucleases, including the C-terminal PCP domain (which is responsible for the self-release of nsp1β from nsp2), a linker domain (LKD) that connects the NTD and the PCP domain, and a C-terminal extension (CTE) that binds to and is stabilized by the putative substrate binding site of the PCPβ domain. Combined with the reported nuclear localization of this protein, these results shed light on the self-processing mode and precise biological function of nsp1β and thus offer a multitarget template for future drug discovery.

Structures of human mitofusin 1 provide insight into mitochondrial tethering

Mitofusin 1 (MFN1) mediates mitochondrial fusion, but the mechanisms involved are unclear. Qi et al. present the crystal structures of a minimal GTPase domain of human MFN1, which suggest that MFN1 tethers apposing membranes through nucleotide-dependent dimerization.

Structural basis for mechanochemical role of Arabidopsis thaliana dynamin-related protein in membrane fission

Dear Editor, Dynamins and dynamin-related proteins (DRPs) constitute a large superfamily of GTPases throughout animal, plant, and bacteria and play essential roles in core cellular processes (Praefcke and McMahon, 2004). Plant specific dynamin-related subfamilies share essential functions with those in mammalian cell, e.g. clarthrinmediated endocytosis and fission of mitochondria; yet they also play unique functional roles in plant cells (Hong et al., 2003; Chen et al., 2011; Xue et al., 2011) (Supplementary Figure S1). Key features of dynamin members, including large molecular size, high basal GTP hydrolysis, and self-assembly into filamentous helices, distinguish them from other classical signaling and regulatory GTPases (Praefcke and McMahon, 2004). Dynamins are known to play a dual-role in clathrinmediated endocytosis, in which the basal activity is necessary for early endocytic events and an assembly-stimulated activity is required in later stages of membrane fission (Sever et al., 1999). A mechanochemical model was presented focusing on dynamin in triggering the vesicle scission stimulated by GTP hydrolysis. In such a model, two distinct mechanisms, i.e. ‘pinchase’ and ‘poppase’, were proposed based on GTP hydrolysis induced dynamin vesiculation on liposomes (Sweitzer and Hinshaw, 1998). Differences between the two possible mechanisms focus on tightening the vesicle neck by dynamin oligomer to ‘pinching off’ the vesicle or a length-wise extension of the dynamin super helix to ‘popping off’’ of the vesicle mechanochemically (Praefcke and McMahon, 2004). To clarify the assembling process and working mechanism of dynamin members in plant cell cytokinetic processes, we initiate the functional and structural investigation on Arabidopsis thaliana dynaminrelated protein 1A (AtDRP1A). GED of dynamin members is known to be directly associated with the GTPase domain (Chappie et al., 2009) and plays a crucial role in either assembly-stimulated or basal GTPase activity. We therefore engineered a 40-kDa AtDRP1A variant containing the GTPase domain and C-terminal segment of GED (CGED, residues 585–606) fused by a flexible linker (Figure 1A) (named as AtDRP1A GG hereafter) as suggested in human dynamin (Chappie et al., 2009). The purified AtDRP1A GG protein maintained a GTPase activity with Km value of 563 mM and kcat of 0.11 min 21 (Supplementary Figure S2), which is in the range of the dynamin family (Praefcke and McMahon, 2004) and suggests that this variant represents the basic catalytic machinery of AtDRP1A. Substrate-dependent oligomerization is known to play a central regulatory role in a number of G proteins (Gasper et al., 2009) including the activity of dynamin’s GTPase domain (Chappie et al., 2010). AtDRP1A GG existed in a monomeric form in the presence of either GDP alone or nonhydrolysable GTP analogues as well as in the absence of additional nucleotides (Supplementary Figure S3A), while mainly existed in a dimeric form when it was incubated with GDP supplemented with NaF and AlCl3 (Supplementary Figure S3A). A hexagonal and a monoclinic crystal form were subsequently obtained with AtDRP1A GG both in dimeric state (Figure 1B). In both crystal forms, GTPase domain presented a canonical structure of GTPase family and two adjacent GTPase cores associated symmetrically with each other (Supplementary Figure S3B). Among the interacting residues, NQDLATSD AIK, named as the ‘trans stabilizing loop’ (Chappie et al., 2010), played a major role in AtDRP1A GG dimerization (Supplementary Figure S3C). Particularly, D186 interacted with G65 thus participating in P-loop stabilization. The side chain of D217 and K218 in the conserved G4 (TKID) motif contributed to intradimer interaction through contacting with the residues in a ‘dynamin specific loop’ (Chappie et al., 2010). More importantly, the hydrophilic side chain of D217 formed in trans interactions with bound GDP molecule as well as G4 motif of the symmetry mate (Supplementary Figure S3D). Although D186A and D217A mutants both forfeit the substrate-dependent dimer formation (Supplementary Figure S4), D186A showed no detectable influence on basal GTPase activity, while D217A significantly reduced GTPase activity (Figure 1C). These observations were consistent with the structural result that D186 is merely involved in protein–protein interaction while D217 also participated in stabilizing GTP molecule. These results reveal that, like in mammalian dynamins (Chappie et al., 2010), oligomerization of AtDRP1A is essential for its stimulated GTP hydrolysis but not the basal GTPase activity. Significantly, distinct differences on ligand binding, active site conformations, and orientations of bundle signaling element (BSE) were observed between the two crystal forms. Whereas the GDP molecule was identified in both crystal forms, AlF4 , resembling the g-phosphate group in the GTP hydrolysis transition status, Mg2+ and Na+, which are crucial for GTP hydrolysis, were only observed in the monoclinic crystal form (Figure 1D). As a consequence of 3.6 Å back bond 378 | Journal of Molecular Cell Biology (2011), 3, 378–381 doi:10.1093/jmcb/mjr032

Structural basis and functional analysis of the SARS coronavirus nsp14–nsp10 complex

Significance Proofreading exonucleases contributing to replication fidelity in DNA viruses and cellular organisms are well known; however, proofreading in RNA viruses was unknown until recently. Coronavirus nonstructural protein 14 (nsp14) has been shown to function as a proofreading exoribonuclease. Additionally, nsp14 shows (guanine-N7) methyl transferase activity for viral mRNA capping. Both roles are important for viral replication and transcription. Here, we report the structures of severe acute respiratory syndrome-coronavirus nsp14 in complex with its activator nonstructural protein 10 (nsp10) and functional ligands. Structural observations coupled with mutagenesis and functional assays provide a better understanding of the function of nsp14. Furthermore, the structures of the nsp14–nsp10 complex demonstrate several unique niches that could be targeted for development of potent antiviral drugs. Nonstructural protein 14 (nsp14) of coronaviruses (CoV) is important for viral replication and transcription. The N-terminal exoribonuclease (ExoN) domain plays a proofreading role for prevention of lethal mutagenesis, and the C-terminal domain functions as a (guanine-N7) methyl transferase (N7-MTase) for mRNA capping. The molecular basis of both these functions is unknown. Here, we describe crystal structures of severe acute respiratory syndrome (SARS)-CoV nsp14 in complex with its activator nonstructural protein10 (nsp10) and functional ligands. One molecule of nsp10 interacts with ExoN of nsp14 to stabilize it and stimulate its activity. Although the catalytic core of nsp14 ExoN is reminiscent of proofreading exonucleases, the presence of two zinc fingers sets it apart from homologs. Mutagenesis studies indicate that both these zinc fingers are essential for the function of nsp14. We show that a DEEDh (the five catalytic amino acids) motif drives nucleotide excision. The N7-MTase domain exhibits a noncanonical MTase fold with a rare β-sheet insertion and a peripheral zinc finger. The cap-precursor guanosine-P3-adenosine-5′,5′-triphosphate and S-adenosyl methionine bind in proximity in a highly constricted pocket between two β-sheets to accomplish methyl transfer. Our studies provide the first glimpses, to our knowledge, into the architecture of the nsp14–nsp10 complex involved in RNA viral proofreading.

Structures of the yeast dynamin-like GTPase Sey1p provide insight into homotypic ER fusion

The crystal structures of the N-terminal cytosolic domain of Sey1p shed light on the mechanism of Sey1p-mediated ER membrane fusion.

Assessment of BAK1 activity in different plant receptor-like kinase complexes by quantitative profiling of phosphorylation patterns

UNLABELLED Plant receptor-like kinases (RLKs) constitute a large family of receptors coordinating developmental programs with adaptation to environmental stresses including immune defenses. BRI1-ASSOCIATED KINASE 1 (BAK1), a member of the plant RLK family, forms receptor complexes with multiple RLK proteins including BRI1, FLS2, EFR and BIK1 to regulate responses to growth hormones or PAMPs. RLK activation and signal initiation involve protein complex formation and phosphorylation/dephosphorylation between BAK1 and its interacting partners. To gain new insight into how phosphorylation contributes to BAK1-mediated signaling specificity, we first mapped the phosphorylation patterns of BAK1 associated with different RLK partners (BRI1, FLS2, EFR and BIK1). Quantitative phospho-pattern profiling by label-free mass spectrometry revealed that differential phosphorylation patterns of RLK partners resulted from altered BAK1 phosphorylation status. More interestingly, the study of two BAK1 mutants (T450A and C408Y) both showing severe defect in immune defense yet normal growth phenotype suggested that varied phosphorylation patterns of RLK partners by BAK1 could be the molecular basis for selective regulation of multiple BAK1-dependent pathways. Taken together, this phospho-pattern profiling strategy allowed for explicit assessment of BAK1 kinase activity in different RLK complexes, which would facilitate elucidation of BAK1 diverse functions in plant development, defense, and adaptation. BIOLOGICAL SIGNIFICANCE BAK1 is a functionally important co-receptor known to interact with different receptor-like kinases (RLKs) to coordinate plant development and immune defenses. Our study first mapped the phosphorylation patterns of BAK1 associated with four RLK partners (BRI1, FLS2, EFR and BIK1), and further revealed that differential phosphorylation patterns of multiple RLK partners resulted from altered BAK1 phosphorylation status. More interestingly, the study of two BAK1 mutants suggested that varied phosphorylation patterns of RLK partners by BAK1 could be the basis for selective regulation of signaling pathways. Taken together, this phospho-pattern profiling strategy allowed for explicit assessment of BAK1 kinase activity in different RLK complexes, which would facilitate elucidation of BAK1 diverse functions in plant development, defense, and adaptation.

Crystal structure of the novel di-nucleotide cyclase from Vibrio cholerae (DncV) responsible for synthesizing a hybrid cyclic GMP-AMP

Crystal structure of the novel di-nucleotide cyclase from Vibrio cholerae (DncV) responsible for synthesizing a hybrid cyclic GMP-AMP