Molecular basis of anti-CRISPR operon repression by Aca10

Lee, So Yeon; Birkholz, Nils; Fineran, Peter C.; Park, Hyun Ho

doi:10.1093/nar/gkac656

Cited by 6 publications

(3 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although various Acrs inhibit CRISPR-Cas activity in monomeric form, previous studies have shown that the dimeric form of Acrs is often critical for their activity ( 29 , 53 , 54 ). Similarly, some Aca proteins have been shown to function as dimers ( 31 , 33 , 55 ). Given the possible dimeric form of AcrIF24 as judged by our SEC experiment, we used multi-angle light scattering (MALS) to confirm the stoichiometry by determining the absolute molecular mass of AcrIF24 in solution.…”

Section: Resultsmentioning

confidence: 99%

Molecular basis of dual anti-CRISPR and auto-regulatory functions of AcrIF24

Kim

Lee

Birkholz

et al. 2022

Nucleic Acids Research

Self Cite

View full text Add to dashboard Cite

CRISPR-Cas systems are adaptive immune systems in bacteria and archaea that provide resistance against phages and other mobile genetic elements. To fight against CRISPR-Cas systems, phages and archaeal viruses encode anti-CRISPR (Acr) proteins that inhibit CRISPR-Cas systems. The expression of acr genes is controlled by anti-CRISPR-associated (Aca) proteins encoded within acr-aca operons. AcrIF24 is a recently identified Acr that inhibits the type I-F CRISPR-Cas system. Interestingly, AcrIF24 was predicted to be a dual-function Acr and Aca. Here, we elucidated the crystal structure of AcrIF24 from Pseudomonas aeruginosa and identified its operator sequence within the regulated acr-aca operon promoter. The structure of AcrIF24 has a novel domain composition, with wing, head and body domains. The body domain is responsible for recognition of promoter DNA for Aca regulatory activity. We also revealed that AcrIF24 directly bound to type I-F Cascade, specifically to Cas7 via its head domain as part of its Acr mechanism. Our results provide new molecular insights into the mechanism of a dual functional Acr-Aca protein.

show abstract

Section: Resultsmentioning

confidence: 99%

Molecular basis of dual anti-CRISPR and auto-regulatory functions of AcrIF24

Kim

Lee

Birkholz

et al. 2022

Nucleic Acids Research

Self Cite

View full text Add to dashboard Cite

show abstract

“…To generate the VEGF 165 /aptamer complexes, we utilized the HDOCK docking program [40,41] by utilizing all combinations of the conformations described in the previous section. In fact, HDOCK has been extensively adopted recently to analyze protein-RNA and protein-DNA interactions [42][43][44], and it has shown excellent performance [45] in the community-wide Critical Assessment of Prediction of Interactions (CAPRI) [46]. To further verify its applicability to our systems, we conducted a docking experiment between VEGF 121 and domains 2 and 3 (D23) of VEGFR-2 with it.…”

Section: Ensemble Docking Of Aptamers With Vegf 165mentioning

confidence: 99%

An Ensemble Docking Approach for Analyzing and Designing Aptamer Heterodimers Targeting VEGF165

Go,

Kalathingal,

Rhee

2024

IJMS

View full text Add to dashboard Cite

Vascular endothelial growth factor 165 (VEGF165) is a prominent isoform of the VEGF-A protein that plays a crucial role in various angiogenesis-related diseases. It is homodimeric, and each of its monomers is composed of two domains connected by a flexible linker. DNA aptamers, which have emerged as potent therapeutic molecules for many proteins with high specificity and affinity, can also work for VEGF165. A DNA aptamer heterodimer composed of monomers of V7t1 and del5-1 connected by a flexible linker (V7t1:del5-1) exhibits a greater binding affinity with VEGF165 compared to either of the two monomers alone. Although the structure of the complex formed between the aptamer heterodimer and VEGF165 is unknown due to the highly flexible linkers, gaining structural information will still be valuable for future developments. Toward this end of accessing structural information, we adopt an ensemble docking approach here. We first obtain an ensemble of structures for both VEGF165 and the aptamer heterodimer by considering both small- and large-scale motions. We then proceed through an extraction process based on ensemble docking, molecular dynamics simulations, and binding free energy calculations to predict the structures of the VEGF165/V7t1:del5-1 complex. Through the same procedures, we reach a new aptamer heterodimer that bears a locked nucleic acid-modified counterpart of V7t1, namely RNV66:del5-1, which also binds well with VEGF165. We apply the same protocol to the monomeric units V7t1, RNV66, and del5-1 to target VEGF165. We observe that V7t1:del5-1 and RNV66:del5-1 show higher binding affinities with VEGF165 than any of the monomers, consistent with experiments that support the notion that aptamer heterodimers are more effective anti-VEGF165 aptamers than monomeric aptamers. Among the five different aptamers studied here, the newly designed RNV66:del5-1 shows the highest binding affinity with VEGF165. We expect that our ensemble docking approach can help in de novo designs of homo/heterodimeric anti-angiogenic drugs to target the homodimeric VEGF165.

show abstract

“…For instance, AcrIC9 from Rhodobacter capsulatus is the most recently identified AcrIC family member that inhibits the type IC CRISPR-Cas system (Gussow et al, 2020). The expression of many acr genes is controlled by putative transcriptional regulators called anti-CRISPR-associated proteins (Acas) (Birkholz et al, 2019;Stanley et al, 2019), which are thus indirect regulators of CRISPR-Cas systems (Lee et al, 2022;Malone et al, 2021). Because Acrs and Acas regulate the activity of CRISPR-Cas systems, they are considered to be promising tools for controlling gene editing using CRISPR-Cas (Marino et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

The structure of AcrIC9 revealing the putative inhibitory mechanism of AcrIC9 against the type IC CRISPR–Cas system

Kang,

Kim,

Lee

et al. 2023

IUCrJ

Self Cite

View full text Add to dashboard Cite

CRISPR–Cas systems are known to be part of the bacterial adaptive immune system that provides resistance against intruders such as viruses, phages and other mobile genetic elements. To combat this bacterial defense mechanism, phages encode inhibitors called Acrs (anti-CRISPR proteins) that can suppress them. AcrIC9 is the most recently identified member of the AcrIC family that inhibits the type IC CRISPR–Cas system. Here, the crystal structure of AcrIC9 from Rhodobacter capsulatus is reported, which comprises a novel fold made of three central antiparallel β-strands surrounded by three α-helixes, a structure that has not been detected before. It is also shown that AcrIC9 can form a dimer via disulfide bonds generated by the Cys69 residue. Finally, it is revealed that AcrIC9 directly binds to the type IC cascade. Analysis and comparison of its structure with structural homologs indicate that AcrIC9 belongs to DNA-mimic Acrs that directly bind to the cascade complex and hinder the target DNA from binding to the cascade.

show abstract

Molecular basis of anti-CRISPR operon repression by Aca10

Cited by 6 publications

References 42 publications

Molecular basis of dual anti-CRISPR and auto-regulatory functions of AcrIF24

Molecular basis of dual anti-CRISPR and auto-regulatory functions of AcrIF24

An Ensemble Docking Approach for Analyzing and Designing Aptamer Heterodimers Targeting VEGF165

The structure of AcrIC9 revealing the putative inhibitory mechanism of AcrIC9 against the type IC CRISPR–Cas system

Contact Info

Product

Resources

About