Discovery and insight into the unique tailoring of the paeninodin lasso peptide from paenibacillus dendritiformis C454

Lasso peptides, such as microcin J25, BI-32169, lariatin and capistruin, are a structurally unique and pharmacologically relevant class of RiPPs (ribosomally synthesized and posttranslationally modified peptides) natural products. Compared with other intensively modified RiPPs, such as lantibiotics, lasso peptides only have a unique knotted topology in which the tail of the peptide is threaded through an N-terminal macrolactam ring and trapped by steric hindrance of bulky side chains stabilizing the entropically disfavored lasso structure. Except for this unusual knot structure, further posttranslational modifications on lasso peptides are very rare. Besides, lasso peptides have so far only been isolated from Proteo- and Actinobacterial sources. In this thesis, the lasso gene cluster from the Firmicute P. dendritiformis was investigated. Paeninodin, a new lasso peptide with an unusual phosphorylation at the side chain of the last serine was discovered by expression of this cluster in a heterologous host. The Paeninodin lasso peptide was isolated from a culture pellet. Mass spectrometric, carboxypeptidase Y assays and IM-MS studies proved paeninodin to be a new representative of lasso peptides. Morever, the biosynthetic pathway of modified lasso peptide was delineated through in vivo and in vitro studies. The kinase turned out to be a novel lasso peptide precursor kinase with wide substrate specificity. These results provide a way for the generation of novel lasso peptide analogs and, thereby, would facilitate lasso peptide engineering in the future

Nanoreactor Design Based on Self-Assembling Protein Nanocages

Self-assembling proteins that form diverse architectures are widely used in material science and nanobiotechnology. One class belongs to protein nanocages, which are compartments with nanosized internal spaces. Because of the precise nanoscale structures, proteinaceous compartments are ideal materials for use as general platforms to create distinct microenvironments within confined cellular environments. This spatial organization strategy brings several advantages including the protection of catalyst cargo, faster turnover rates, and avoiding side reactions. Inspired by diverse molecular machines in nature, bioengineers have developed a variety of self-assembling supramolecular protein cages for use as biosynthetic nanoreactors that mimic natural systems. In this mini-review, we summarize current progress and ongoing efforts creating self-assembling protein based nanoreactors and their use in biocatalysis and synthetic biology. We also highlight the prospects for future research on these versatile nanomaterials

Discovery and insight into the unique tailoring of the paeninodin lasso peptide from paenibacillus dendritiformis C454

Lasso peptides, such as microcin J25, BI-32169, lariatin and capistruin, are a structurally unique and pharmacologically relevant class of RiPPs (ribosomally synthesized and posttranslationally modified peptides) natural products. Compared with other intensively modified RiPPs, such as lantibiotics, lasso peptides only have a unique knotted topology in which the tail of the peptide is threaded through an N-terminal macrolactam ring and trapped by steric hindrance of bulky side chains stabilizing the entropically disfavored lasso structure. Except for this unusual knot structure, further posttranslational modifications on lasso peptides are very rare. Besides, lasso peptides have so far only been isolated from Proteo- and Actinobacterial sources. In this thesis, the lasso gene cluster from the Firmicute P. dendritiformis was investigated. Paeninodin, a new lasso peptide with an unusual phosphorylation at the side chain of the last serine was discovered by expression of this cluster in a heterologous host. The Paeninodin lasso peptide was isolated from a culture pellet. Mass spectrometric, carboxypeptidase Y assays and IM-MS studies proved paeninodin to be a new representative of lasso peptides. Morever, the biosynthetic pathway of modified lasso peptide was delineated through in vivo and in vitro studies. The kinase turned out to be a novel lasso peptide precursor kinase with wide substrate specificity. These results provide a way for the generation of novel lasso peptide analogs and, thereby, would facilitate lasso peptide engineering in the future

MOESM1 of Construction of an organelle-like nanodevice via supramolecular self-assembly for robust biocatalysts

Additional file 1: Table S1. Primers for cloning, mutagenesis of the MhIHL and for construction of the nanoreactor. Fig. S1. Schematic and sequence of artificially fused protein open reading frame for γ-lactamase nanoreactor. Fig. S2. Size-exclusion chromatography (SEC) of the free and encapsulated (+)-γ-lactamases. Fig. S3. TEM structures of free γ-lactamase and encapsulated γ-lactamase self-assembled in vitro. Fig. S4. AFM structures of free γ-lactamase and encapsulated γ-lactamase self-assembled in vitro. Fig. S5. Structures of empty protein dodecahedron formed by the engineered ketohydroxyglutarate aldolase from Thermotoga maritima. Fig. S6. Michaelis–Menten plot used to determine the Km and kcat values. Fig. S7. Optimal temperature for free γ-lactamase and encapsulated γ-lactamase. Fig. S8. Optimal pH for free γ-lactamase and encapsulated γ-lactamase. Fig. S9. Comparison of the stability of the free and encapsulated γ-lactamase in presence of protease. Fig. S10. The Lineweaver–Burk double reciprocal plot for different concentrations of (+)-γ-lactam

Synthesis of MOFs and Their Composite Structures through Sacrificial-Template Strategy

Exemplified by chemical conversion of ZnO nanostructures into zeolitic imidazolate framework-8 (ZIF-8) nanostructures, a sacrificial-template method has been demonstrated for the synthesis of metal–organic frameworks (MOFs) and their composite structures which may not be attainable by other methods. Their properties were investigated and the formation mechanism of ZIF-8 nanostructures was discussed. This method shows the potential of the formation of various-shaped MOFs and their composite nanostructures and will broad the applications of MOFs and their derivatives

The ring residue proline 8 is crucial for the thermal stability of the lasso peptide caulosegnin II

Lasso peptides are fascinating natural products with a unique structural fold that can exhibit tremendous thermal stability.</p

Discovery and characterization of Rubrinodin provide clues into the evolution of Lasso peptides

Lasso peptides are unique natural products that comprise a class of ribosomally synthesized and post-translationally modified peptides. Their defining three-dimensional structure is a lariat knot, in which the C-terminal tail is threaded through a macrolactam ring formed between the N-terminal amino group and an Asp or Glu side chain (i.e., an isopeptide bond). Recent genome mining strategies have revealed various types of lasso peptide biosynthetic gene clusters and have thus redefined the known chemical space of lasso peptides. To date, over 20 different types of these gene clusters have been discovered, including several different clades from Proteobacteria. Despite the diverse architectures of these gene clusters, which may or may not encode various tailoring enzymes, most currently known lasso peptides are synthesized by two discrete clades defined by the presence of an ATP-binding cassette transporter or its absence and (sometimes) concurrent appearance of an isopeptidase, raising questions about their evolutionary history. Herein, we discovered and characterized the lasso peptide rubrinodin, which is assembled by a gene cluster encoding both an ATP-binding cassette transporter and an isopeptidase. Our bioinformatics analyses of this and other representative cluster types provided new clues into the evolutionary history of lasso peptides. Furthermore, our structural and biochemical investigations of rubrinodin permitted the conversion of this thermolabile lasso peptide into a more thermostable scaffold

Effects of initial mist conditions on simulation accuracy of humidity distribution in an environmental chamber

This paper firstly presents experimental study of humidity distribution in an environmental chamber at different inlet temperatures and humidity. Experiment results show that, under fixed inlet temperature condition, the uniformity of the relative humidity (RH) in the chamber is better when the inlet RH is higher. It is mainly because of the coupled heat and mass transfer occurred between water droplets and air. In practice, direct measurement of humidity distribution in a chamber is infeasible and CFD simulation is a possible way to predict the humidity distribution. However, the use of CFD tools for predicting humidity field is rarely reported. Boundary and initial mist conditions are critical to the accuracy and computation efficiency of CFD simulation. This study then presents a CFD simulation method of the humidity distribution inside chamber using FLUENT based on some assumptions and simplification. Moreover, droplet sizes were measured by laser diffraction analyzer. The simplified droplet size distribution is used as initial condition for CFD simulation. Experiment results and CFD simulation results are compared and it can be concluded that the CFD simulation method can achieve satisfactory accuracy with reasonable computation load