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Current Opinion in Chemical Biology Jun 2020Engineered proteins are revolutionizing immunotherapy, but advances are still needed to harness their full potential. Traditional protein engineering methods use... (Review)
Review
Engineered proteins are revolutionizing immunotherapy, but advances are still needed to harness their full potential. Traditional protein engineering methods use naturally existing proteins as a starting point, and therefore, are intrinsically limited to small alterations of a protein's natural structure and function. Conversely, computational de novo protein design is free of such limitation, and can produce a virtually infinite number of novel protein sequences, folds, and functions. Recently, we used de novo protein engineering to create Neoleukin-2/15 (Neo-2/15), a protein mimetic of the function of both interleukin-2 (IL-2) and interleukin-15 (IL-15). To our knowledge, Neo-2/15 is the first de novo protein with immunotherapeutic activity, and in murine cancer models, it has demonstrated enhanced therapeutic potency and reduced toxicity compared to IL-2. De novo protein design is already showcasing its tremendous potential for driving the next wave of protein-based therapeutics that are explicitly engineered to treat disease.
Topics: Amino Acid Sequence; Animals; Immunotherapy; Interleukin-15; Interleukin-2; Mice; Models, Molecular; Neoplasms; Neoplasms, Experimental; Protein Binding; Protein Conformation; Protein Engineering; Structure-Activity Relationship
PubMed: 32371023
DOI: 10.1016/j.cbpa.2020.02.002 -
Journal of Molecular Evolution Aug 2022"De novo" genes evolve from previously non-genic DNA. This strikes many of us as remarkable, because it seems extraordinarily unlikely that random sequence would produce... (Review)
Review
"De novo" genes evolve from previously non-genic DNA. This strikes many of us as remarkable, because it seems extraordinarily unlikely that random sequence would produce a functional gene. How is this possible? In this two-part review, I first summarize what is known about the origins and molecular functions of the small number of de novo genes for which such information is available. I then speculate on what these examples may tell us about how de novo genes manage to emerge despite what seem like enormous opposing odds.
Topics: Evolution, Molecular
PubMed: 35451603
DOI: 10.1007/s00239-022-10055-3 -
Methods in Molecular Biology (Clifton,... 2022Development of computer-aided de novo design methods to discover novel compounds in a speedy manner to treat human diseases has been of interest to drug discovery... (Review)
Review
Development of computer-aided de novo design methods to discover novel compounds in a speedy manner to treat human diseases has been of interest to drug discovery scientists for the past three decades. In the beginning, the efforts were mostly concentrated to generate molecules that fit the active site of the target protein by sequential building of a molecule atom-by-atom and/or group-by-group while exploring all possible conformations to optimize binding interactions with the target protein. In recent years, deep learning approaches are applied to generate molecules that are iteratively optimized against a binding hypothesis (to optimize potency) and predictive models of drug-likeness (to optimize properties). Synthesizability of molecules generated by these de novo methods remains a challenge. This review will focus on the recent development of synthetic planning methods that are suitable for enhancing synthesizability of molecules designed by de novo methods.
Topics: Artificial Intelligence; Drug Discovery; Humans; Proteins
PubMed: 34731479
DOI: 10.1007/978-1-0716-1787-8_17 -
International Journal of Molecular... Feb 2023Food enzymes have an important role in the improvement of certain food characteristics, such as texture improvement, elimination of toxins and allergens, production of... (Review)
Review
Food enzymes have an important role in the improvement of certain food characteristics, such as texture improvement, elimination of toxins and allergens, production of carbohydrates, enhancing flavor/appearance characteristics. Recently, along with the development of artificial meats, food enzymes have been employed to achieve more diverse functions, especially in converting non-edible biomass to delicious foods. Reported food enzyme modifications for specific applications have highlighted the significance of enzyme engineering. However, using direct evolution or rational design showed inherent limitations due to the mutation rates, which made it difficult to satisfy the stability or specific activity needs for certain applications. Generating functional enzymes using de novo design, which highly assembles naturally existing enzymes, provides potential solutions for screening desired enzymes. Here, we describe the functions and applications of food enzymes to introduce the need for food enzymes engineering. To illustrate the possibilities of using de novo design for generating diverse functional proteins, we reviewed protein modelling and de novo design methods and their implementations. The future directions for adding structural data for de novo design model training, acquiring diversified training data, and investigating the relationship between enzyme-substrate binding and activity were highlighted as challenges to overcome for the de novo design of food enzymes.
Topics: Protein Engineering; Functional Food; Proteins; Enzymes
PubMed: 36835238
DOI: 10.3390/ijms24043827 -
Genes Sep 2021The genetic etiology of congenital diaphragmatic hernia (CDH), a common and severe birth defect, is still incompletely understood. Chromosomal aneuploidies, copy number... (Review)
Review
The genetic etiology of congenital diaphragmatic hernia (CDH), a common and severe birth defect, is still incompletely understood. Chromosomal aneuploidies, copy number variations (CNVs), and variants in a large panel of CDH-associated genes, both and inherited, have been described. Due to impaired reproductive fitness, especially of syndromic CDH patients, and still significant mortality rates, the contribution of variants to the genetic background of CDH is assumed to be high. This assumption is supported by the relatively low recurrence rate among siblings. Advantages in high-throughput genome-wide genotyping and sequencing methods have recently facilitated the detection of variants in CDH. This review gives an overview of the known disease-causing variants in CDH patients.
Topics: Aneuploidy; Chromosome Aberrations; DNA Copy Number Variations; Hernias, Diaphragmatic, Congenital; Humans; Mutation
PubMed: 34573387
DOI: 10.3390/genes12091405 -
Methods in Molecular Biology (Clifton,... 2022In the latest years, the application of deep generative models to suggest virtual compounds is becoming a new and powerful tool in drug discovery projects. The idea... (Review)
Review
In the latest years, the application of deep generative models to suggest virtual compounds is becoming a new and powerful tool in drug discovery projects. The idea behind this review is to offer an updated view on de novo design approaches based on artificial intelligent (AI) algorithms, with a particular focus on ligand-based methods. We start this review by reporting a brief overview of the most relevant de novo design approaches developed before the use of AI techniques. We then describe the nowadays most common neural network architectures employed in ligand-based de novo design, together with an up-to-date list of more than 100 deep generative models found in the literature (2017-2020). In order to show how deep generative approaches are applied into drug discovery context, we report all the now available studies in which generated compounds have been synthetized and their biological activity tested. Finally, we discuss what we envisage as beneficial future directions for further application of deep generative models in de novo drug design.
Topics: Artificial Intelligence; Deep Learning; Drug Design; Ligands; Neural Networks, Computer
PubMed: 34731474
DOI: 10.1007/978-1-0716-1787-8_12 -
Neurosurgical Focus Jul 2022Seizures are the second most common presenting symptom of brain arteriovenous malformations (bAVMs) after hemorrhage. Risk factors for preoperative seizures and...
OBJECTIVE
Seizures are the second most common presenting symptom of brain arteriovenous malformations (bAVMs) after hemorrhage. Risk factors for preoperative seizures and subsequent seizure control outcomes have been well studied. There is a paucity of literature on postoperative, de novo seizures in initially seizure-naïve patients who undergo resection. Whereas this entity has been documented after craniotomy for a wide variety of neurosurgically treated pathologies including tumors, trauma, and aneurysms, de novo seizures after bAVM resection are poorly studied. Given the debilitating nature of epilepsy, the purpose of this study was to elucidate the incidence and risk factors associated with de novo epilepsy after bAVM resection.
METHODS
A retrospective review of patients who underwent resection of a bAVM over a 15-year period was performed. Patients who did not present with seizure were included, and the primary outcome was de novo epilepsy (i.e., a seizure disorder that only manifested after surgery). Demographic, clinical, and radiographic characteristics were compared between patients with and without postoperative epilepsy. Subgroup analysis was conducted on the ruptured bAVMs.
RESULTS
From a cohort of 198 patients who underwent resection of a bAVM during the study period, 111 supratentorial ruptured and unruptured bAVMs that did not present with seizure were included. Twenty-one patients (19%) developed de novo epilepsy. One-year cumulative rates of developing de novo epilepsy were 9% for the overall cohort and 8.5% for the cohort with ruptured bAVMs. There were no significant differences between the epilepsy and no-epilepsy groups overall; however, the de novo epilepsy group was younger in the cohort with ruptured bAVMs (28.7 ± 11.7 vs 35.1 ± 19.9 years; p = 0.04). The mean time between resection and first seizure was 26.0 ± 40.4 months, with the longest time being 14 years. Subgroup analysis of the ruptured and endovascular embolization cohorts did not reveal any significant differences. Of the patients who developed poorly controlled epilepsy (defined as Engel class III-IV), all had a history of hemorrhage and half had bAVMs located in the temporal lobe.
CONCLUSIONS
De novo epilepsy after bAVM resection occurs at an annual cumulative risk of 9%, with potentially long-term onset. Younger age may be a risk factor in patients who present with rupture. The development of poorly controlled epilepsy may be associated with temporal lobe location and a delay between hemorrhage and resection.
Topics: Brain; Embolization, Therapeutic; Epilepsy; Humans; Intracranial Arteriovenous Malformations; Retrospective Studies; Seizures; Treatment Outcome
PubMed: 35901740
DOI: 10.3171/2022.4.FOCUS2288 -
Genome Medicine May 2023Long-read sequencing (LRS) techniques have been very successful in identifying structural variants (SVs). However, the high error rate of LRS made the detection of small...
BACKGROUND
Long-read sequencing (LRS) techniques have been very successful in identifying structural variants (SVs). However, the high error rate of LRS made the detection of small variants (substitutions and short indels < 20 bp) more challenging. The introduction of PacBio HiFi sequencing makes LRS also suited for detecting small variation. Here we evaluate the ability of HiFi reads to detect de novo mutations (DNMs) of all types, which are technically challenging variant types and a major cause of sporadic, severe, early-onset disease.
METHODS
We sequenced the genomes of eight parent-child trios using high coverage PacBio HiFi LRS (~ 30-fold coverage) and Illumina short-read sequencing (SRS) (~ 50-fold coverage). De novo substitutions, small indels, short tandem repeats (STRs) and SVs were called in both datasets and compared to each other to assess the accuracy of HiFi LRS. In addition, we determined the parent-of-origin of the small DNMs using phasing.
RESULTS
We identified a total of 672 and 859 de novo substitutions/indels, 28 and 126 de novo STRs, and 24 and 1 de novo SVs in LRS and SRS respectively. For the small variants, there was a 92 and 85% concordance between the platforms. For the STRs and SVs, the concordance was 3.6 and 0.8%, and 4 and 100% respectively. We successfully validated 27/54 LRS-unique small variants, of which 11 (41%) were confirmed as true de novo events. For the SRS-unique small variants, we validated 42/133 DNMs and 8 (19%) were confirmed as true de novo event. Validation of 18 LRS-unique de novo STR calls confirmed none of the repeat expansions as true DNM. Confirmation of the 23 LRS-unique SVs was possible for 19 candidate SVs of which 10 (52.6%) were true de novo events. Furthermore, we were able to assign 96% of DNMs to their parental allele with LRS data, as opposed to just 20% with SRS data.
CONCLUSIONS
HiFi LRS can now produce the most comprehensive variant dataset obtainable by a single technology in a single laboratory, allowing accurate calling of substitutions, indels, STRs and SVs. The accuracy even allows sensitive calling of DNMs on all variant levels, and also allows for phasing, which helps to distinguish true positive from false positive DNMs.
Topics: Humans; High-Throughput Nucleotide Sequencing; Alleles; INDEL Mutation; Microsatellite Repeats
PubMed: 37158973
DOI: 10.1186/s13073-023-01183-6 -
Current Opinion in Structural Biology Jun 2021Comparative evolutionary genomics has revealed that novel protein coding genes can emerge randomly from non-coding DNA. While most of the myriad of transcripts which... (Review)
Review
Comparative evolutionary genomics has revealed that novel protein coding genes can emerge randomly from non-coding DNA. While most of the myriad of transcripts which continuously emerge vanish rapidly, some attain regulatory regions, become translated and survive. More surprisingly, sequence properties of de novo proteins are almost indistinguishable from randomly obtained sequences, yet de novo proteins may gain functions and integrate into eukaryotic cellular networks quite easily. We here discuss current knowledge on de novo proteins, their structures, functions and evolution. Since the existence of de novo proteins seems at odds with decade-long attempts to construct proteins with novel structures and functions from scratch, we suggest that a better understanding of de novo protein evolution may fuel new strategies for protein design.
Topics: Evolution, Molecular; Genomics; Proteins
PubMed: 33567396
DOI: 10.1016/j.sbi.2020.11.010 -
Andrology Jan 2021Are de novo mutations in the human genome associated with male infertility?
STUDY QUESTION
Are de novo mutations in the human genome associated with male infertility?
SUMMARY ANSWER
We identified de novo mutations in five candidate genes: SEMA5A, NEURL4, BRD2, CD1D, and CD63.
WHAT IS KNOWN ALREADY
Epidemiological and genetic studies have consistently indicated contribution of genetic factors to the etiology of male infertility, suggesting that more than 1500 genes are involved in spermatogenesis.
STUDY DESIGN, SIZE, DURATION
First, we searched for de novo mutations in patients with idiopathic azoospermia with whole-exome sequencing (WES). To evaluate the potential functional impact of de novo identified mutations, we analyzed their expression differences on independent testis samples with normal and impaired spermatogenesis. In the next step, we tested additional group of azoospermic patients for mutations in identified genes with de novo mutations. In addition to the analysis of de novo mutations in patients with idiopathic azoospermia, we considered other models of inheritance and searched for candidate genes harboring rare maternally inherited variants and biallelic autosomal and X-chromosome hemizygous variants.
PARTICIPANTS/MATERIALS, SETTING, METHODS
We performed WES in 13 infertile males with idiopathic azoospermia and their parents. Potential functional impact of de novo identified mutations was evaluated by global gene expression profiling on 20 independent testis samples. To replicate the results, we performed WES in further 16 independent azoospermic males, which were screened for the variants in the same genes. Library preparation was performed with Nextera Coding Exome Capture Kit (Illumina), with subsequent sequencing on Illumina HiSeq 2500 platform.
MAIN RESULTS AND THE ROLE OF CHANCE
We identified 11 de novo mutations in 10 genes of which 5 were considered potentially associated with azoospermia: SEMA5A, NEURL4, BRD2, CD1D, and CD63. All candidate genes showed significant differential expression in testis samples composed of patients with severely impaired and normal spermatogenesis. Additionally, we identified rare, potentially pathogenic mutations in the genes previously implicated in male infertility-a maternally inherited heterozygous frameshift variant in FKBPL gene and inframe deletion in UPF2 gene, homozygous frameshift variant in CLCA4 gene, and a heterozygous missense variant NR0B1 gene, which represent promising candidates for further clinical implication.
LIMITATIONS OF THE STUDY, REASONS FOR CAUTION
We provided limited functional support for involvement of de novo identified genes in pathogenesis of male infertility, based on expression analysis. Additionally, the sample size was limited.
WIDER IMPLICATIONS OF THE FINDINGS
We provide support that de novo mutations might contribute to male infertility and propose five genes as potentially implicated in its pathogenesis.
Topics: Humans; Infertility, Male; Male; Pilot Projects; Point Mutation; Exome Sequencing
PubMed: 32860660
DOI: 10.1111/andr.12897