SmartBac, a new baculovirus system for large protein complex production

Zhai, Yujia; Zhang, Danyang; Yu, Leiye; Sun, Fei; Sun, Fei

doi:10.1016/j.yjsbx.2019.100003

Cited by 17 publications

(17 citation statements)

References 54 publications

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“…Here, the CATCHR domains of Sec5 and Sec8 of subcomplex I form large interfaces with subcomplex II, and Sec6 connects both subcomplexes like an arch. A preliminary 3D reconstruction of human exocyst from negatives stain EM yielded a particle of similar overall shape and size, suggesting that exocyst architecture is conserved …”

Section: Catchr Multisubunit Tethersmentioning

confidence: 99%

Structure of membrane tethers and their role in fusion

Ungermann

Kümmel

2019

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Vesicular transport between different membrane compartments is a key process in cell biology required for the exchange of material and information. The complex machinery that executes the formation and delivery of transport vesicles has been intensively studied and yielded a comprehensive view of the molecular principles that underlie the budding and fusion process. Tethering also represents an essential step in each trafficking pathway. It is mediated by Rab GTPases in concert with so-called tethering factors, which constitute a structurally diverse family of proteins that share a similar role in promoting vesicular transport. By simultaneously binding to proteins and/or lipids on incoming vesicles and the target compartment, tethers are thought to bridge donor and acceptor membrane. They thus provide specificity while also promoting fusion. However, how tethering works at a mechanistic level is still elusive.We here discuss the recent advances in the structural and biochemical characterization of tethering complexes that provide novel insight on how these factors might contribute the efficiency of fusion. K E Y W O R D S CATCHR, golgin, membrane fusion, Rab GTPases, Sec1/Munc18, SNARE, tethering, vesicular transport 1 | INTRODUCTION Compartmentalization of eukaryotic cells into organelles is the prerequisite for the formation of chemically distinct environments that allow the implementation of specialized functions. Communication and the exchange of substances between different membrane compartments, however, is also essential for proper function of cells. In the endomembrane system, comprising the secretory pathway and the endolysosomal system, membrane vesicles have long been established as transport carriers that fulfill this function. 1 Trafficking can be dissected into distinct steps: at the donor membrane, small GTPases like Arf, Sar1 or Arf-like (Arl) and coat proteins, which assemble into vesicle coat complexes, orchestrate cargo sorting and concentration followed by vesicle budding and scission. Vesicles are then transported along cytoskeletal filaments with the help of motor proteins. At the target membrane, incoming vesicles are recruited by Rab GTPases and tethering factors. Rabs are markers of organelle identity and conserved part of the fusion machinery. 2,3 Common to all small GTPases, Rabs cycle between an inactive GDP-bound and an active GTP-bound form, and only the latter form can interact with effectors that mediate downstream functions of Rabs. Tethering factors from different classes are evolutionarily unrelated proteins and structurally divers, but share a function as Rab effectors and are thought to bridge two opposing membranes. After tethering, SNAREs (soluble N-ethylmaleimide sensitive factor attachment protein receptor) on each membrane are necessary. 4 SNARE proteins drive fusion, but function of SNAREs requires S/M (Sec1/Munc18) family proteins that act as chaperones of SNARE assembly and the NSF (N-ethylmaleimide sensitive factor) ATPase that with the adaptor protein α-SNAP disasse...

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Section: Catchr Multisubunit Tethersmentioning

confidence: 99%

Structure of membrane tethers and their role in fusion

Ungermann

Kümmel

2019

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“…For the human MPC2, an additional EGFP (enhanced green fluorescent protein) tag was added before the His-tag. The use of the C-terminal EGFP tag was widespread for membrane proteins [ 49 , 50 ], including the purification of large multiprotein complexes [ 51 , 52 ], and was particularly useful in detecting inducible expression levels, solubility, stability, and the cellular compartmentalization of recombinant MPC [ 31 ].…”

Section: Contextual Discussionmentioning

confidence: 99%

Insights on the Quest for the Structure–Function Relationship of the Mitochondrial Pyruvate Carrier

Quesñay

Pollock

Nagampalli

et al. 2020

Biology

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The molecular identity of the mitochondrial pyruvate carrier (MPC) was presented in 2012, forty years after the active transport of cytosolic pyruvate into the mitochondrial matrix was first demonstrated. An impressive amount of in vivo and in vitro studies has since revealed an unexpected interplay between one, two, or even three protein subunits defining different functional MPC assemblies in a metabolic-specific context. These have clear implications in cell homeostasis and disease, and on the development of future therapies. Despite intensive efforts by different research groups using state-of-the-art computational tools and experimental techniques, MPCs’ structure-based mechanism remains elusive. Here, we review the current state of knowledge concerning MPCs’ molecular structures by examining both earlier and recent studies and presenting novel data to identify the regulatory, structural, and core transport activities to each of the known MPC subunits. We also discuss the potential application of cryogenic electron microscopy (cryo-EM) studies of MPC reconstituted into nanodiscs of synthetic copolymers for solving human MPC2.

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“…This study found that cell lysis could be delayed and that recombinant protein yields could be increased by using cell lines constitutively expressing vankyrin or vankyrin-encoding baculovirus vectors (Steele et al, 2017). SmartBac, a new baculovirus system, was developed for large protein complex production (Zhai et al, 2019). The FlexiBAC protein expression system was also developed for the production of both cytosolic proteins and secreted proteins that require proteolytic maturation.…”

Section: Insect Cellsmentioning

confidence: 99%

Recent Developments in Bioprocessing of Recombinant Proteins: Expression Hosts and Process Development

Tripathi

Shrivastava

2019

Front. Bioeng. Biotechnol.

397

243

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Infectious diseases, along with cancers, are among the main causes of death among humans worldwide. The production of therapeutic proteins for treating diseases at large scale for millions of individuals is one of the essential needs of mankind. Recent progress in the area of recombinant DNA technologies has paved the way to producing recombinant proteins that can be used as therapeutics, vaccines, and diagnostic reagents. Recombinant proteins for these applications are mainly produced using prokaryotic and eukaryotic expression host systems such as mammalian cells, bacteria, yeast, insect cells, and transgenic plants at laboratory scale as well as in large-scale settings. The development of efficient bioprocessing strategies is crucial for industrial production of recombinant proteins of therapeutic and prophylactic importance. Recently, advances have been made in the various areas of bioprocessing and are being utilized to develop effective processes for producing recombinant proteins. These include the use of high-throughput devices for effective bioprocess optimization and of disposable systems, continuous upstream processing, continuous chromatography, integrated continuous bioprocessing, Quality by Design, and process analytical technologies to achieve quality product with higher yield. This review summarizes recent developments in the bioprocessing of recombinant proteins, including in various expression systems, bioprocess development, and the upstream and downstream processing of recombinant proteins.

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SmartBac, a new baculovirus system for large protein complex production

Cited by 17 publications

References 54 publications

Structure of membrane tethers and their role in fusion

Structure of membrane tethers and their role in fusion

Insights on the Quest for the Structure–Function Relationship of the Mitochondrial Pyruvate Carrier

Recent Developments in Bioprocessing of Recombinant Proteins: Expression Hosts and Process Development

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