Article
Biochemistry & Molecular Biology
Rafael Molina, Anne Louise Gron Jensen, Javier Marchena-Hurtado, Blanca Lopez-Mendez, Stefano Stella, Guillermo Montoya
Summary: Type III CRISPR-Cas effector systems detect foreign RNA and trigger cell dormancy or death. Standalone ring nucleases downregulate the immune response by degrading cA, thus controlling the cell antiviral state.
NUCLEIC ACIDS RESEARCH
(2021)
Article
Chemistry, Multidisciplinary
Olivia Luige, Partha Pratim Bose, Rouven Stulz, Peter Steunenberg, Omar Brun, Shalini Andersson, Merita Murtola, Roger Stromberg
Summary: The study introduces Zn2+-dependent dimethyl-dipyridophenazine PNA conjugates as efficient RNA cleaving artificial enzymes, showing site-specific RNA cleavage with 10 minute half-lives and cleaving clinically relevant RNA models.
CHEMICAL COMMUNICATIONS
(2021)
Article
Biochemistry & Molecular Biology
Xiaoman Jiang, Yang Liu, Qi Liu, Lixin Ma
Summary: This study discovered three active prokaryotic Argonautes (pAgos) with about 50% sequence identity to KmAgo, which have typical DNA-guided DNA target cleavage ability. Among them, RsuAgo from Rummeliibacillus suwonensis was mainly explored for its ability to cleave both DNA and RNA targets, especially with short guide DNAs in the range of 16-20 nt. RsuAgo-mediated RNA target cleavage showed a preference for a guide strand with a 5'-terminal A residue. Furthermore, RsuAgo was found to be able to cleave double-stranded DNA in a low-salt buffer at 37 degrees C, providing a new tool for DNA and RNA manipulation at moderate temperatures.
Article
Biochemistry & Molecular Biology
Benjamin Ober-Reynolds, Winston R. Becker, Karina Jouravleva, Samson M. Jolly, Phillip D. Zamore, William J. Greenleaf
Summary: Studies have shown that TtAgo uses biophysical adaptations similar to those of eukaryotic Argonautes for rapid association but requires more extensive complementarity to achieve high-affinity target binding. The cleavage rates of TtAgo vary widely based on the DNA guide, suggesting that only a subset of guides cleaves targets on physiologically relevant timescales.
Review
Biochemistry & Molecular Biology
Yu Chen, Zhifeng Zeng, Qunxin She, Wenyuan Han
Summary: CRISPR-Cas and pAgo are defense systems in prokaryotes that use nucleic acid (NA) guides to recognize and cleave invading NA targets. However, they also utilize the abortive infection (Abi) strategy to activate toxic effectors and kill infected cells, preventing the spread of invaders. This review summarizes the mechanisms of CRISPR-Cas and pAgo systems and highlights their crucial roles in the microbial arms race against invaders.
TRENDS IN MICROBIOLOGY
(2023)
Article
Multidisciplinary Sciences
Qichen Yuan, Xue Gao
Summary: This study introduces a novel method for multiplex gene editing in human cells using CRISPR array architecture and RNA polymerase III promoter, enabling simultaneous editing of multiple disease-related genomic loci.
NATURE COMMUNICATIONS
(2022)
Article
Multidisciplinary Sciences
Ildar Gainetdinov, Joel Vega-Badillo, Katharine Cecchini, Ayca Bagci, Cansu Colpan, Dipayan De, Shannon Bailey, Amena Arif, Pei-Hsuan Wu, Ian J. MacRae, Phillip D. Zamore
Summary: In eukaryotes, small RNA guides direct the Argonaute protein family to regulate gene expression and defend the genome. PIWI proteins, a clade unique to animals, use PIWI-interacting RNAs to repress transposon transcripts. Unlike AGO proteins, PIWI proteins can efficiently cleave transcripts that are only partially paired to their guides. PIWI slicing tolerates mismatches to any target nucleotide, making it better equipped to target rapidly diverging transposons without new small RNA guides.
Article
Biochemical Research Methods
Danial Pourjafar-Dehkordi, Martin Zacharias
Summary: Argonaute proteins in combination with short microRNAs target mRNA for translation inhibition or degradation. The study reveals that binding of guide and target induces domain movements in Argonaute, with 15 or 16 base pairs required for activation. This provides a mechanistic explanation for the observed activation process.
PLOS COMPUTATIONAL BIOLOGY
(2021)
Article
Biochemistry & Molecular Biology
Wenlong Zhu, Stuart McQuarrie, Sabine Gruschow, Stephen A. McMahon, Shirley Graham, Tracey M. Gloster, Malcolm F. White
Summary: A newly discovered CARF family effector named Can2 is found to provide effective immunity against plasmid transformation and bacteriophage infection in Escherichia coli by being activated by cyclic tetra-adenylate (cA(4)) and displaying both DNase and RNase activity. The structure of Can2 in complex with cA(4) reveals a potential mechanism for the cA(4)-mediated activation of the enzyme, where an active site cleft is exposed upon activator binding. These findings enhance our understanding of type III CRISPR cOA signaling and effector function.
NUCLEIC ACIDS RESEARCH
(2021)
Article
Biochemistry & Molecular Biology
Rafael Molina, Ricardo Garcia-Martin, Blanca Lopez-Mendez, Anne Louise Gron Jensen, J. Rafael Ciges-Tomas, Javier Marchena-Hurtado, Stefano Stella, Guillermo Montoya
Summary: Standalone ring nucleases are CRISPR ancillary proteins that downregulate the immune response of Type III CRISPR-Cas systems. A study on Sulfolobus islandicus genome identified two genes coding for different length polypeptides with this function. The analysis of one of these polypeptides revealed its structure and the key role it plays in the cleavage process.
NUCLEIC ACIDS RESEARCH
(2022)
Review
Chemistry, Multidisciplinary
Huimin Li, Yi Xie, Fumin Chen, Huiwen Bai, Leshan Xiu, Xiaonong Zhou, Xiaokui Guo, Qinqin Hu, Kun Yin
Summary: Rapid and accurate molecular diagnosis is crucial for precision medicine, food safety, and environmental monitoring. CRISPR/Cas-based detection has become an effective tool for molecular diagnosis due to its outstanding advantages. However, existing methods typically require pre-amplification, and the development of target amplification-free CRISPR/Cas-based detection still faces challenges.
CHEMICAL SOCIETY REVIEWS
(2023)
Article
Chemistry, Multidisciplinary
Yangdao Wei, Zhiqing Yang, Chengli Zong, Buhua Wang, Xiaolin Ge, Xiao Tan, Xin Liu, Zhenzhen Tao, Peng Wang, Chunxin Ma, Yi Wan, Jinghong Li
Summary: Cas14a1 exhibits a novel capacity where RNA can trigger its cleavage of DNA. The RNA-activated detection platform based on Cas14a1 shows high specificity for target RNAs with point mutations and can efficiently detect pathogenic nucleic acids, serving as a new tool for nucleic acid diagnostics.
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
(2021)
Article
Biochemistry & Molecular Biology
Ya-Jing Shi, Min Duan, Jun-Mei Ding, Fan-Qi Wang, Li-Li Bi, Cai-Xiang Zhang, Yi-Zhou Zhang, Jun-Yi Duan, An-Hui Huang, Xin-Lin Lei, Hao Yin, Ying Zhang
Summary: A Cas9 protein (Alicyclobacillus tengchongensis) from a thermophilic bacterium has been found to regulate PAM recognition through DNA topology. Structural analysis reveals that AtCas9 has a unique loop motif that interacts with the DNA major groove to modulate PAM recognition. Experimental results demonstrate the near-PAMless editing capability of AtCas9 in E. coli and its broad PAM preference for efficient editing in mammalian cells.
Article
Biochemistry & Molecular Biology
Rachelle R. Q. Lee, Wei Yuan Cher, Jinge Wang, Yujie Chen, Eunyoung Chae
Summary: MINORg is an offline gRNA design tool that generates the smallest combination of gRNA to cover desired targets in multiple non-reference genomes. By utilizing sequence homology and screening multiple genetic backgrounds, MINORg aims to reduce workload and create reusable gRNA panels. Practical application of MINORg was demonstrated by knocking out homologous genes in Arabidopsis thaliana and implementing a PCR-free modular cloning system. The source code is freely available at https://github.com/rlrq/MINORg.
NUCLEIC ACIDS RESEARCH
(2023)
Article
Biochemistry & Molecular Biology
So Young Bak, Youngri Jung, Jinho Park, Keewon Sung, Hyeon-Ki Jang, Sangsu Bae, Seong Keun Kim
Summary: This study utilized single-molecule FRET assay to investigate the dynamics of DNA:RNA heteroduplexation for various engineered Cas9 variants, revealing a strong correlation between their conformational structure and nuclease activity. Quantitative analysis showed that these engineered Cas9 variants exhibit a more rapid decrease in cleavage-competent state with increasing base-pair mismatch, leading to enhanced target specificity. The critical specificity enhancement step was found to be activated during DNA:RNA heteroduplexation for some variants, highlighting the importance of conformational dynamics in Cas9 target specificity.
NUCLEIC ACIDS RESEARCH
(2021)
Article
Biochemistry & Molecular Biology
Sajal Maity, Madhuri Hande, Tuomas Lonnberg
Article
Biochemistry & Molecular Biology
Asmo Aro-Heinila, Tuomas Lonnberg, Pasi Virta
Summary: By incorporating a highly nucleobase-discriminating metalated nucleoside analogue into an oligonucleotide molecular beacon, clear binding selectivity was achieved at room temperature, with a preference order of G>T>C>A. This method allows for robust discrimination between different nucleobases and is suitable for the detection of single nucleotide polymorphisms.
Article
Biochemistry & Molecular Biology
Lange Yakubu Saleh, Mikko Ora, Tuomas Lonnberg
Summary: A water-soluble arylmercury complex has been synthesized and its ability to catalyze the cleavage of the phosphodiester linkage of the RNA model compound has been evaluated. The catalytic reaction is linearly dependent on catalyst concentration and shows relatively low affinity for the substrate.
Review
Biochemistry & Molecular Biology
Dattatraya Ukale, Tuomas Lonnberg
Summary: This review summarizes the synthetic efforts towards nucleosides, nucleotides, oligonucleotides, and nucleic acids covalently mercurated at their base moieties, as well as discusses the proposed, realized, and abandoned applications of such compounds. Emphasis is placed on ongoing research areas, particularly the use of Hg-II-mediated base pairing to enhance the hybridization properties of oligonucleotide probes. The review also aims to anticipate potential future applications of organomercury nucleic acids.
Review
Biochemistry & Molecular Biology
Satu Mikkola
Article
Biochemistry & Molecular Biology
Juho Jaakkola, Anu Nieminen, Henri Kivela, Heidi Korhonen, Petri Tahtinen, Satu Mikkola
Summary: The study investigated the alkaline cleavage of sugar nucleotides using H-1 and P-31 NMR, revealing the complex reaction systems and significant effects of conditions on reactivity. The results were consistent with proposed mechanisms, despite the inability to identify individual intermediate species.
NUCLEOSIDES NUCLEOTIDES & NUCLEIC ACIDS
(2021)
Article
Biochemistry & Molecular Biology
Madhuri Hande, Sajal Maity, Tuomas Lonnberg
Summary: The study investigated the effects of alkylpyridine C-nucleoside derivatives or respective palladacycles on short oligodeoxynucleotides, revealing a decrease in duplex stability upon palladation. Competition experiments showed a more pronounced impact at the 3' terminus compared to the 5' terminus. The dependence of UV melting temperature on the identity of canonical nucleobases opposite to modified nucleobases was more significant in the palladacyclic duplexes.
JOURNAL OF INORGANIC BIOCHEMISTRY
(2021)
Article
Chemistry, Organic
Asmo Aro-Heinila, Assi Lepisto, Antti Aarela, Tuomas Antti Lonnberg, Pasi Virta
Summary: A novel organomercury nucleobase analogue was synthesized and incorporated into an oligonucleotide, followed by mercuration. The affinity of the organomercury nucleobase analogue towards complementary strands was studied using UV, CD, and F-19 NMR spectroscopy, revealing distinct affinities for different canonical nucleobases. Results showed that the F-19 resonance signal was sensitive to local environmental changes, providing valuable information on the binding of the mercurated oligonucleotide to complementary strands at varying temperatures.
JOURNAL OF ORGANIC CHEMISTRY
(2022)
Article
Chemistry, Organic
Josefiina Wallin, Tuomas Lonnberg
Summary: The dependence of the hydrolysis rate of a series of N-methoxy-2-phenyl-1,3-oxazinanes on the Hammett substituent constant of the substituent of the phenyl ring was determined. Based on this information, 4-(benzoyloxy)benzaldehyde was selected as a protecting group for a new (2R,3S)-4-(methoxyamino)butane-1,2,3-triol phosphoramidite building block. The yield of the preparation of this building block as well as its coupling in automated oligonucleotide synthesis were greatly improved compared to the method reported previously.
EUROPEAN JOURNAL OF ORGANIC CHEMISTRY
(2022)
Article
Chemistry, Multidisciplinary
Tharun K. Kotammagari, Petri Tahtinen, Tuomas Lonnberg
Summary: Short oligonucleotides containing organometallic moieties were synthesized and studied as hybridization probes for sequences containing thiopyrimidine nucleobases. They showed stable duplex formation with 4-thiothymine and were able to discriminate between 2- and 4-thiothymine based on the melting temperature. The DFT-optimized structures revealed the coordination and bonding patterns between Hg-II ions and sulfur or oxygen substituents.
CHEMISTRY-A EUROPEAN JOURNAL
(2022)
Article
Chemistry, Organic
Maaret Toiviala, Vesa Kleemola, Sajal Maity, Tuomas Lonnberg
Summary: Both alpha and beta anomers of an acetophenone C-nucleoside were synthesized and incorporated in the middle of short oligodeoxynucleotides. The ketone oligonucleotides were converted to N-15-labelled oxime oligonucleotides and cyclopalladated. Comparison of the UV melting profiles suggested formation of a stable Pd(II)-mediated base pair with adenine or thymine as the base pairing partner. The N-15 NMR spectra indicated successful labeling, but the signals diminished below detection limit when hybridized with a complementary strand in a specific configuration.
EUROPEAN JOURNAL OF ORGANIC CHEMISTRY
(2023)
Article
Biochemistry & Molecular Biology
Satu Mikkola, Olga Eguaogie, Anu Nieminen, Patrick F. Conlon, David L. Jakeman, Keith Moore, Ian C. Lane, Joseph S. Vyle
Summary: In this study, novel sulfur and selenium substituted dinucleoside pyrophosphate analogues were prepared using a vibration ball mill and their chemical hydrolysis reactions were investigated. The results showed that the chalcogeno-substitution had a modest effect on the reactivity of dinucleoside pyrophosphates and exhibited promising chemical stability. The hydrolytic desilylation of precursor phosphorochalcogenolate monoesters was also studied and the rate of dephosphorylation depended on the nature of the chalcogen and the pH.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2022)
Article
Biochemistry & Molecular Biology
Lange Yakubu Saleh, Mikko Ora, Tuomas Lonnberg
Summary: Two oligonucleotide conjugates were synthesized with different 5'-terminal organometallic moieties through direct mercuration or oximation. These conjugates exhibited higher catalytic activity as artificial ribonucleases compared to metal-free counterparts and previously reported small molecule organomercury RNA cleaving agent. However, the activity was modest compared to state-of-the-art metal-dependent artificial ribonucleases.
JOURNAL OF INORGANIC BIOCHEMISTRY
(2023)
Article
Chemistry, Multidisciplinary
Mark N. K. Afari, Kasper Nurmi, Pasi Virta, Tuomas Lonnberg
Summary: Hairpin oligodeoxynucleotides with a modified residue in the stem were synthesized and reacted with aromatic aldehydes to produce nucleoside analogues. The reaction equilibrium depends on the aldehyde and nucleobase. The modified residue showed the highest affinity and selectivity with 9-formyl-9-deazaadenine.
Article
Chemistry, Organic
Mark N. K. Afari, Pasi Virta, Tuomas Lonnberg
Summary: In this study, (2R,3S)-4-(methoxyamino)butane-1,2,3-triol was converted into a protected phosphoramidite building block and incorporated into short DNA oligonucleotides, replacing the furanose ring with an oxazinane ring through reaction with aldehydes. This approach demonstrated the feasibility of synthesizing base-modified oligonucleotides using aromatic and aliphatic aldehydes with various functional groups.
ORGANIC & BIOMOLECULAR CHEMISTRY
(2022)