Nature Newsletter

Hello everyone, this issue brings you Nature Communications, a subsidiary of Nature Group, which publishes high-quality research papers in biology, physics, chemistry and other fields. The impact factor in 2020 is 14.9438+0.

1

The cryoelectron microscopic structure of human A2ML 1 illustrates the mechanism of A2M family protease inhibition.

The frozen electron microstructure of human A2ML 1 shows the mechanism of A2M family protease inhibition.

A2ML 1 is a monomeric protease inhibitor, belonging to A2M superfamily of protease inhibitors and complement factors. In this study, the author studied the protease inhibition mechanism of human A2ML 1, and determined the structure of its natural and proteolytic cleavage conformation. The functional inhibitory unit of A2ML 1 is a monomer, which is inhibited by the valence binding of protease (mediated by thioester of A2ML 1). Compared with A2M tetramer capturing protease in two internal chambers formed by four subunits, in the protease-cleaved monomer A2ML 1, disordered regions surround the captured protease and may prevent the substrate from entering. In natural A2ML 1, the decoy region passes through hydrophobic channels, which indicates that disrupting this arrangement by cutting the decoy region will lead to a wide range of conformational changes, which will lead to protease inhibition. Compared with C3/C4, A2M protein superfamily has this mechanism, which can trigger the conformational changes after proteolysis activation.

2

The selective origin of Streptococcus Siglec-like adhesin suggests the receptor adaptation mechanism.

The selective origin of polysaccharide in Streptococcus-like adhesin indicates the receptor adaptation mechanism.

The combination of bacteria and host receptors is the basis of * *. Many streptococci use Siglec-like binding region (SLBR) to adhere to protein-linked carbohydrates expressed on the cell surface. The identified accurate glycan library can determine whether the organism is strictly an organism rather than a pathogen. However, it is not clear what drives the selectivity of receptors. In this study, the author used five representative SLBR and identified receptor binding sites with high sequence and structure. The results show that these regions use chimeras and single amino acid substitutions to control the identity of preferred carbohydrate ligands. The authors further evaluated how the identity of the preferred ligand affects the interaction with glycoprotein receptors in human saliva and plasma samples. Because point mutation can change the preferred human receptor, these studies show how streptococcus adapts to the changes of environmental glycan pool.

three

Computationally designed super-active Cas9 enzyme

Computationally designed Cas9 enzyme with high activity

The ability to change the genome of living cells is the key to understand how genes affect the function of organisms, and it is very important to modify living systems to achieve useful purposes. However, this goal has long been limited by the technical challenges involved in genetic engineering. The latest progress in gene editing has bypassed some of these challenges, but the results are not satisfactory. In this study, the author calculated and designed Cas9 enzyme with significantly higher donor-independent editing activity by using FuncLib. The author used the genetic circuit related to yeast cell survival to quantify Cas9 activity, and found the synergy between engineering fields. These overactive Cas9 variants have played an effective role in mammalian cells and introduced larger and more diverse insertion and deletion pools in the target genome region, which provides a tool for enhancing and expanding the possible application of gene editing based on CRISPR.

four

Modular (de-) construction of complex bacterial phenotype by CRISPR/nCas9-assisted multiple cytidine base editing

Complex bacterial phenotypes were analyzed by CRISPR/nCas9 assistance and polycytidine base editing.

Modular (decomposition) structure

CRISPR/Cas technology constitutes a powerful tool for genome engineering, but their use in non-traditional bacteria depends on host factors or exogenous recombinant enzymes, which limits efficiency and flux. In this study, the author eased these practical limitations by developing a widely applicable tool set for genome engineering of Gram-negative bacteria. This challenge has been solved by customizing CRISPR base editor, which can achieve 90% efficiency by using >: single nucleotide splitting operation (C G T A). In addition, GUIDRNAs processing mediated by Cas6 is integrated into the streamlined scheme of plasmid assembly, which supports multi-base editing and the efficiency is >: 85%. This tool is used to construct and deconstruct the complex phenotype of pseudomonas putida in soil bacteria. One-step engineering of aromatic compounds production phenotype and multi-step deconstruction of complex redox metabolism illustrate the versatility of multi-base editing provided by this toolbox. Therefore, this method overcomes the typical limitations of the previous technology and gives a Gram-negative bacteria engineering scheme that is far away.

five

Using protein Group Constraints to Improve the Yield of Recombinant Yeast Protein by Genome Scale Modeling

Using protein Group to Constrain Genome Scale Modeling to Improve Yeast Recombinant Protein Production

As a cell factory, eukaryotic cells produce and secrete a large number of recombinant drug proteins, including several best-selling drugs at present. Because of the important role and complexity of secretion pathway, it is relatively temporary to increase the yield of recombinant protein through metabolic engineering traditionally. And a more systematic method is needed to generate novel design principles. In this study, the author proposed a genome-scale protein secretion model constrained by protein Group of Saccharomyces cerevisiae, which made it possible to simulate and explain the phenotype caused by limited secretion ability. The author further predicted the overexpression targets of several recombinant proteins by using pcSecYeast model. Many predicted α -amylase production targets were verified by experiments, which proved the application of pcSecYeast as a calculation tool in guiding yeast engineering and improving recombinant protein production.

six

In vivo gene amplification system for high-level expression in Saccharomyces cerevisiae

In vivo gene amplification system with high level expression in Saccharomyces cerevisiae

The bottleneck of metabolic pathway caused by insufficient gene expression level is still a big problem in industrial biological production by using microbial cell factory. Increasing gene dose can overcome these bottlenecks, but the current method has many shortcomings. In this study, the author describes HapAmp, a method that uses haploid defects as an evolutionary force to drive gene amplification in vivo. HapAmp can realize efficient, titratable and stable integration of foreign gene copies, and at most 47 copies can be transferred to yeast genome. Taking metabolic engineering as an example, this method can significantly increase the yield of sesquiterpene neroli oil, monoterpene limonene and tetraterpene lycopene. The titer of limonene was increased by 20 times in a single engineering step, and it was1g L-1in shake flask culture. The author also showed that the yield of heterologous protein in yeast increased significantly. HapAmp is an effective method to quickly unlock the bottleneck of metabolism, which is used for the development of microbial cell factories.

seven

Discovery and characterization of terpene biosynthesis pathway characterized by Diels-alder enzyme forming norbornene

Discovery and characterization of terpene biosynthesis pathway formed by norbornene Diels-alder enzyme

Pericyclic enzymes that catalyze pericyclic reactions form an expanding family of enzymes with biocatalysis. Although more and more pericyclic enzymes have been found, it is surprising that the Diels-Alder cyclization reaction between cyclopentadiene and olefin dienophile produces norbornene, which is one of the best cycloaddition reactions in synthetic chemistry, and there is no corresponding enzymatic reaction so far. In this study, the author reported the discovery of a pathway characterized by norbornene synthase SdnG, which was used to synthesize terpene precursors of the antifungal natural product Sorbus pohuashanensis. The complete reconstruction of sordaricin biosynthesis reveals the simple oxidation strategy used in nature, which transforms the complete hydrocarbon precursor into a highly functional substrate of SdnG for intramolecular Diels-Alder cycloaddition. SdnG produces the norbornene core of sorboside and accelerates the reaction to inhibit the host-mediated redox modification of activated dienophiles. The discovery of this work has expanded the scope of pericyclic enzyme-catalyzed reaction and P450-mediated terpene maturation.

eight

Reasonable modification of sandalwood synthase to adjust the composition ratio of sandalwood oil

Rationally transform sandalwood synthase and adjust the proportion of sandalwood oil components.

Plant essential oil (PEO) is widely used in cosmetics and health care industries. The composition ratio of PEO determines their quality. In the construction of PEO biotechnology platform, it is a challenge to control the proportion of components. In this study, the author explored the catalytic reaction pathways of product hybrids and product-specific sandalwood synthases (SaSSy and SanSyn) through multi-scale simulation. It is found that F44 1 of SanSyn is the key residue that limits the conformational dynamics of the intermediate, so the direct deprotonation of general base T298 mainly produces α-sandalwood. The subsequent mutation of plastic residues produced mutant enzyme SanSynF44 1V, which can produce α-and β-sandalwood. Through the efforts of metabolic engineering, the titer of sandalwood terpene/sandalwood phenol reached 704.2 mg/L, and the composition ratio matched the ISO 35 18:2002 standard very well. This study provides an example for the combination of metabolism and enzyme engineering to build a PEO biotechnology platform with ideal composition ratio.