A novel alkane monooxygenase evolved from a broken piece of ribonucleotide reductase in Geobacillus kaustophilus HTA426 isolated from Mariana Trench.

We have accidentally found that a thermophilic Geobacillus kaustophilus HTA426 is capable of degrading alkanes although it has no alkane oxygenating enzyme genes. Our experimental results revealed that a putative ribonucleotide reductase small subunit GkR2loxI (GK2771) gene encodes a novel heterodinuclear Mn-Fe alkane monooxygenase/hydroxylase. GkR2loxI protein can perform two-electron oxidations similar to homonuclear diiron bacterial multicomponent soluble methane monooxygenases. This finding not only answers a long-standing question about the substrate of the R2lox protein clade, but also expands our understanding of the vast diversity and new evolutionary lineage of the bacterial alkane monooxygenase/hydroxylase family.

Assessment of TET1 gene expression, DNA methylation and H3K27me3 level of its promoter region in eutopic endometrium of women with endometriosis and infertility.

Endometriosis is the cause of infertility. The eutopic endometrium of women with endometriosis showed an aberrant expression pattern of multitude genes. The role of TET1 protein in the pathogenesis of endometriosis and related infertility is not sufficiently known. Further, knowledge on TET1 transcriptional control still remains incomplete. The aim of the study was assessment of TET1 gene expression, DNA methylation and H3K27me3 level of its promoter region in eutopic endometrium of women with endometriosis and infertility. The study included 44 infertile patients with endometriosis (IWE) and 77 infertile (IW) and fertile (FW) patients without endometriosis. The research material was eutopic endometrium. The TET1 mRNA level was analyzed by qPCR. Western blot was used to evaluate the level of TET1 protein. The level of DNA methylation and H3K27me3 level of TET1 gene's promoter region were assessed using HRM and ChIP qPCR, respectively. The level of TET1 expression (TET1 mRNA; TET1 protein level) was lower in IWE during the implantation window (p < 0.001; p = 0.0329). The level of TET1 DNA methylation was higher in the secretory endometrium in mild and advanced IWE (p < 0.004; p < 0.008). H3K27me3 level did not differ between the study groups. The diminished expression of TET1 gene during the secretory phase, may account for the aberrant process of embryonic implantation in infertile endometriosis patients. DNA hypermethylation of TET1 gene is a potential relevant regulator of its expression. H3K27me3 occupancy does not affect the expression of TET1 gene in our study group.

Functional genomics assessment of lytic polysaccharide mono-oxygenase with glycoside hydrolases in Paenibacillus dendritiformis CRN18.

Recently discovered Lytic Polysaccharide Mono-Oxygenase (LPMO) enhances the enzymatic deconstruction of complex polysaccharide by oxidation. The present study demonstrates the agricultural waste hydrolyzing capabilities of Paenibacillus dendritiformis CRN18, which exhibits the enzyme activity of exo-glucanase, ß-glucosidase, ß-glucuronidase, endo-1, 4 ß-xylanases, arabinosidase, and α-galactosidase as 0.1U/ml, 0.3U/ml, 0.09U/ml, 0.1U/ml, 0.05U/ml, and 0.41U/ml, respectively. The genome analysis of strain reveals the presence of four LPMO genes, along with lignocellulolytic genes. The gene structure of LPMO and its phylogenetic analysis shows the evolutionary relatedness with the Bacillus LPMO gene. Gene position of LPMOs in the genome of strains shows the close association of two LPMOs with chitin active enzyme GH18, and the other two are associated with hemicellulases (GH39, GH23). Protein-protein interaction and gene networking of LPMO sheds light on the co-occurrence, neighborhood, and interaction of LPMOs with chitinase and xylanase enzymes. Structural prediction of LPMOs unravels the information of the LPMO's binding site. Although the LPMO has been explored for its oxidative mechanism, a little light has been shed on its gene structure. This study provides insights into the LPMO gene structure in P. dendritiformis CRN18 and its potential in lignocellulose hydrolysis.

Evaluation of the CRISPR/Cas9 Genetic Constructs in Efficient Disruption of Porcine Genes for Xenotransplantation Purposes Along with an Assessment of the Off-Target Mutation Formation.

The increasing life expectancy of humans has led to an increase in the number of patients with chronic diseases and organ failure. However, the imbalance between the supply and the demand for human organs is a serious problem in modern transplantology. One of many solutions to overcome this problem is the use of xenotransplantation. The domestic pig (Sus scrofa domestica) is currently considered as the most suitable for human organ procurement. However, there are discrepancies between pigs and humans that lead to the creation of immunological barriers preventing the direct xenograft. The introduction of appropriate modifications to the pig genome to prevent xenograft rejection is crucial in xenotransplantation studies. In this study, porcine GGTA1, CMAH, ß4GalNT2, vWF, ASGR1 genes were selected to introduce genetic modifications. The evaluation of three selected gRNAs within each gene was obtained, which enabled the selection of the best site for efficient introduction of changes. Modifications were examined after nucleofection of porcine primary kidney fibroblasts with CRISPR/Cas9 system genetic constructs, followed by the tracking of indels by decomposition (TIDE) analysis. In addition, off-target analysis was carried out for selected best gRNAs using the TIDE tool, which is new in the research conducted so far and shows the utility of this tool in these studies.

Characterization of inheritance and preliminary biochemical mechanisms of spirotetramat resistance in Phenacoccus solenopsis Tinsley: An economic pest from Pakistan.

Phenacoccus solenopsis is an economically important insect pest of different agronomic and horticultural field crops. In Pakistan, the cotton crop was severely attacked by P. solenopsis during 2007 and since then a varied group of insecticides are used by farmers to manage this pest. As a result, insecticide resistance has become a barrier in control of P. solenopsis. The current study was designed to explore the basics of genetics, realized heritability and possible genetic mechanisms of resistance against spirotetramat in P. solenopsis. Before selection, the wild population (Wild-Pop) showed 5.97-fold resistance when compared with lab-reared susceptible strain (Susceptible Lab-Pop). The P. solenopsis was selected with spirotetramat to 21 generations, called Spiro-SEL Pop, which showed 463.21-fold resistance as compared with the Susceptible Lab-Pop. The values of LC50 for F1 (Spiro-SEL Pop ♂ × Susceptible Lab-Pop ♀) and F1 (Spiro-SEL Pop ♀ × Susceptible Lab-Pop ♂) populations were statistically similar and values of dominance level were 0.42 and 0.54, respectively. Reciprocal crosses between Susceptible Lab-Pop and Spiro-SEL Pop showed that resistance was of autosomal in nature with incomplete dominant traits. According to the fit test, monogenic model estimation of the number of genes, which are responsible for the development of spirotetramat resistance in a population of P. solenopsis, showed that multiple genes are involved in controlling the resistance levels in tested strains of P. solenopsis. The value of heritability for resistance against spirotetramat was 0.13 in P. solenopsis. Our results suggested the presence of a metabolic-based resistance mechanism associated with the monooxygenases in P. solenopsis, while testing the synergism mechanism. These results will provide the baseline to design an effective control strategy to manage P. solenopsis in the field.

The discovery of novel LPMO families with a new Hidden Markov model.

BACKGROUND: Renewable biopolymers, such as cellulose, starch and chitin are highly resistance to enzymatic degradation. Therefore, there is a need to upgrade current degradation processes by including novel enzymes. Lytic polysaccharide mono-oxygenases (LPMOs) can disrupt recalcitrant biopolymers, thereby enhancing hydrolysis by conventional enzymes. However, novel LPMO families are difficult to identify using existing methods. Therefore, we developed a novel profile Hidden Markov model (HMM) and used it to mine genomes of ascomycetous fungi for novel LPMOs. RESULTS: We constructed a structural alignment and verified that the alignment was correct. In the alignment we identified several known conserved features, such as the histidine brace and the N/Q/E-X-F/Y motif and previously unidentified conserved proline and glycine residues. These residues are distal from the active site, suggesting a role in structure rather than activity. The multiple protein alignment was subsequently used to build a profile Hidden Markov model. This model was initially tested on manually curated datasets and proved to be both sensitive (no false negatives) and specific (no false positives). In some of the genomes analyzed we identified a yet unknown LPMO family. This new family is mostly confined to the phyla of Ascomycota and Basidiomycota and the class of Oomycota. Genomic clustering indicated that at least some members might be involved in the degradation of ß-glucans, while transcriptomic data suggested that others are possibly involved in the degradation of pectin. CONCLUSIONS: The newly developed profile hidden Markov Model was successfully used to mine fungal genomes for a novel family of LPMOs. However, the model is not limited to bacterial and fungal genomes. This is illustrated by the fact that the model was also able to identify another new LPMO family in Drosophila melanogaster. Furthermore, the Hidden Markov model was used to verify the more distant blast hits from the new fungal family of LPMOs, which belong to the Bivalves, Stony corals and Sea anemones. So this Hidden Markov model (Additional file 3) will help the broader scientific community in identifying other yet unknown LPMOs.

Safety assessment of dicamba mono-oxygenases that confer dicamba tolerance to various crops.

Dicamba tolerant (DT) soybean, cotton and maize were developed through constitutive expression of dicamba mono-oxygenase (DMO) in chloroplasts. DMO expressed in three DT crops exhibit 91.6-97.1% amino acid sequence identity to wild type DMO. All DMO forms maintain the characteristics of Rieske oxygenases that have a history of safe use. Additionally, they are all functionally similar in vivo since the three DT crops are all tolerant to dicamba treatment. None of these DMO sequences were found to have similarity to any known allergens or toxins. Herein, to further understand the safety of these DMO variants, a weight of evidence approach was employed. Each purified DMO protein was found to be completely deactivated in vitro by heating at temperatures 55 °C and above, and all were completely digested within 30 s or 5 min by pepsin and pancreatin, respectively. Mice orally dosed with each of these DMO proteins showed no adverse effects as evidenced by analysis of body weight gain, food consumption and clinical observations. Therefore, the weight of evidence from all these protein safety studies support the conclusion that the various forms of DMO proteins introduced into DT soybean, cotton and maize are safe for food and feed consumption, and the small amino acid sequence differences outside the active site of DMO do not raise any additional safety concerns.

A multiplex assay to detect variations in the CYP2C9, VKORC1, CYP4F2 and APOE genes involved in acenocoumarol metabolism.

OBJECTIVES: We have developed a genotyping system to determine the alleles of genes related to interindividual variability in acenocoumarol dosage requirements. This genotyping system is intended for routine clinical use and therefore it is essential that it be simple, fast and inexpensive. DESIGN AND METHODS: We developed a PCR multiplex SNaPshot reaction that targets 6 SNPs (single nucleotide polymorphisms) in CYP2C9, CYP4F2, VKORC1 and APOE genes, which are associated with acenocoumarol dose requirements. RESULTS: We tested the multiplex in 152 samples and found it to be 100% concordant with the results of other methods. CONCLUSIONS: We successfully produced a reliable multiplex system for simultaneously typing 6 SNPs. This system may be used as a model for accurate, simple and inexpensive genotyping of SNPs related to dose requirements. This information allows the prediction of drug efficiency in patients prior to treatment with acenocoumarol and the prevention of adverse drug reactions.

Accuracy assessment of pharmacogenetic algorithms for warfarin dose prediction in Chinese patients.

A few warfarin pharmacogenetic dosing algorithms have been proposed,based on multiethnic or homogeneous populations, to estimate warfarin therapeutic doses. However, it remains to be proven that which algorithm is accurate in predicting warfarin dose in Chinese people. We selected eight warfarin dose predictive pharmacogenetic algorithms and retrospectively assessed the predictive accuracy of each algorithm in a total of 368 eligible outpatients by comparing the actual stable therapeutic dose to the dose predicted by the algorithm. Our results showed that a high level of performance was demonstrated by three algorithms,Gage et al., Anderson et al., and Wu et al., having a similar performance in coefficient of determination (R2) and percentage of patients predicted dose within 20% of actual dose. The Gage et al. algorithm had the lowest mean absolute error (MAE). These results indicated that the algorithm by Gage et al. provided a more accurate prediction than did the others,which suggests that this pharmacogenetic algorithm might be used in clinical practice to guide rational administration of warfarin.

New allele-specific real-time PCR system for warfarin dose genotyping equipped with an automatic interpretative function that allows rapid, accurate, and user-friendly reporting in clinical laboratories.

INTRODUCTION: The optimal dose of the oral anticoagulant warfarin varies with polymorphisms of the vitamin K epoxide reductase complex subunit 1 (VKORC1) and cytochrome P450 2C9 (CYP2C9) genes. A fast and reliable method of warfarin dose adjustment is required to prevent serious hemorrhagic or thrombotic complications. The aim of this study is to develop and validate a new warfarin dose genotyping system with an automatic interpretation function. MATERIALS AND METHODS: Four VKORC1 and two CYP2C9 SNPs were genotyped by real-time PCR using allele-specific primers and probes. Multiple reactions that included internal positive controls were performed in each well, and an automatic interpretative algorithm was developed. This system was validated using 82 clinical specimens previously genotyped by PCR-direct sequencing. The analytical time of the method was calculated. RESULTS: No interference was observed when multiple samples were included in each reaction, with all internal positive control reactions being successful. In the genotyping algorithm, Ct differences <2 and ≥2 identified heterozygotes and homozygotes, respectively. All results obtained were concordant with those of the reference method. The overall analytical time for assay of 12 specimens was around 3 hours. CONCLUSION: This rapid, accurate, and user-friendly genotyping system improves the efficacy and safety of anticoagulation therapy in clinical practice.

Benefits of pharmacogenomics in drug development-earlier launch of drugs and less adverse events.

Currently, pharmaceutical companies are reluctant to introduce pharmacogenomics (PGx) in their practice, since cost-benefit of PGx is obscure and methodology to use PGx in drug development has not been fully established yet. The purpose of this study is to investigate advantages obtained by introducing PGx in clinical trials. Particularly, taking Warfarin as an example, we investigate benefits of Enrichment effect that raises response rate of the drug by PGx. When response rate is raised by only 5%, cost of a clinical trial can be reduced to about 40% of a conventional clinical trial. Furthermore, since period necessary for a trial also can be reduced, development period can be shortened by about 750 days. In summary, PGx enables earlier launch of a drug with less cost, representing benefit to pharmaceutical companies, patients and public as a whole.

Will there be a role for genotyping in warfarin therapy?

PURPOSE OF REVIEW: In North America warfarin is the current standard for oral anticoagulation therapy in the treatment and/or prophylaxis of different thrombotic conditions. In daily clinical practice a significant proportion of patients on long-term warfarin therapy fail to stabilize within their target therapeutic range leading to a resultant increased risk of thromboembolism or bleeding. Various authors and agencies advocate the role of genetic testing to guide warfarin dosing. RECENT FINDINGS: Evidence regarding the clinical efficacy and cost-effectiveness of genotype-based warfarin dosing has been conflicting, although some recent studies have suggested a potential benefit in certain subgroups. SUMMARY: More evidence is needed before the wide adoption of genotype-based warfarin dosing. Future studies should be designed to address outcomes such as major bleeding or recurrent thrombosis, and allow economic evaluations.

The impact of genetic polymorphisms and patient characteristics on warfarin dose requirements: a cross-sectional study in Iran.

BACKGROUND: Warfarin is the most commonly prescribed oral anticoagulant drug for prophylaxis and treatment of venous and arterial thromboembolic disorders. Its anticoagulant effect is widely variable between patients because of pharmacodynamic, pharmacokinetic, and pharmacogenetic factors. OBJECTIVE: This study was conducted to identify the associations between demographic characteristics, warfarin maintenance dose, and genetic polymorphisms of cytochrome P450 (CYP) 2C19, CYP2C9, and vitamin K epoxide reductase complex subunit 1 (VKORC1). METHODS: This study was conducted from April 2005 to April 2008 at 3 warfarin clinics affiliated with Shiraz University of Medical Sciences. Blood samples were collected from patients with stable warfarin maintenance dose and a stable target international normalized ratio of 2 to 3. Patients who had a condition (including use of an interacting medication) affecting the metabolism of warfarin were excluded. CYP2C9, CYP2C19, and VKORC1 genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism. The associations between demographic characteristics (eg, age, sex, body surface area, weight, height), genetic factors, and maintenance warfarin dose were examined by multiple linear regression. The probability of F as a criterion for removal of a variable from the multiple linear regression was set at 0.1. RESULTS: One hundred patients were enrolled in the study; complete data were available for 55, who were included in the regression analysis. Among this smaller group, the mean (SD) age was 53 (11) years (range, 25-80 years) and mean weight was 72 (15) kg (range, 42-125 kg); the mean warfarin dose was 27.2 (13.4) mg/week. The allelic frequencies of CYP2C9*2 and CYP2C9*3 were 27% and 9%, respectively. The allelic frequencies of CYP2C19*2 and CYP2C19*3 were 11% and 1%, respectively. Fifteen percent of our patients carried a VKORC1 genotype GG, whereas the AA and GA genotypes were seen in 18% and 58% of patients, respectively. Multiple linear regression analysis found that sex (P = 0.045), height (P = 0.024), age (P = 0.081), and VKORC1 (P = 0.004) and CYP2C9 (P = 0.011) polymorphism had significant influence on the maintenance dose of warfarin. They were associated with 41.3% of the variability in warfarin maintenance dose requirement. VKORC1 polymorphism (partial R(2) = 20.3%) and height (partial R(2) = 20.3%) had the greatest effects on warfarin maintenance dose requirement. CONCLUSION: Among the demographic and genetic factors evaluated in these Iranian patients, sex, height, age, CYP2C9, and VKORC1 had significant effects on warfarin maintenance dose requirements.

Implementing genotype-guided antithrombotic therapy.

Genotyping has the potential to improve the efficacy and safety of major antithrombotic drugs. For warfarin, the stable maintenance dose varies from 1-10 mg/day. The VKORC1 -1639G>A allele and the CYP2C9*2 and *3 alleles (cumulative frequency: 90% in Asians, 65% in Europeans and 20% in Africans), explain 45% of response variability in European and 30% in African populations. The large clinical trials COAG and EU-PACT will define the extent to which pharmacogenetic dosing affects the safety and efficacy of warfarin and coumarin derivatives. The platelet inhibitor clopidogrel requires activation by the CYP2C19 enzyme. CYP2C19*2 and *3 alleles (cumulative frequency: 20-50%) produce null enzyme activity, and their presence attenuates platelet inhibition and increases cardiovascular events. The US FDA-mandated drug labeling recognizes the relevance of genotyping in the selection and dosing of both warfarin and clopidogrel.

Clinical assessment incorporating a personal genome.

BACKGROUND: The cost of genomic information has fallen steeply, but the clinical translation of genetic risk estimates remains unclear. We aimed to undertake an integrated analysis of a complete human genome in a clinical context. METHODS: We assessed a patient with a family history of vascular disease and early sudden death. Clinical assessment included analysis of this patient's full genome sequence, risk prediction for coronary artery disease, screening for causes of sudden cardiac death, and genetic counselling. Genetic analysis included the development of novel methods for the integration of whole genome and clinical risk. Disease and risk analysis focused on prediction of genetic risk of variants associated with mendelian disease, recognised drug responses, and pathogenicity for novel variants. We queried disease-specific mutation databases and pharmacogenomics databases to identify genes and mutations with known associations with disease and drug response. We estimated post-test probabilities of disease by applying likelihood ratios derived from integration of multiple common variants to age-appropriate and sex-appropriate pre-test probabilities. We also accounted for gene-environment interactions and conditionally dependent risks. FINDINGS: Analysis of 2.6 million single nucleotide polymorphisms and 752 copy number variations showed increased genetic risk for myocardial infarction, type 2 diabetes, and some cancers. We discovered rare variants in three genes that are clinically associated with sudden cardiac death-TMEM43, DSP, and MYBPC3. A variant in LPA was consistent with a family history of coronary artery disease. The patient had a heterozygous null mutation in CYP2C19 suggesting probable clopidogrel resistance, several variants associated with a positive response to lipid-lowering therapy, and variants in CYP4F2 and VKORC1 that suggest he might have a low initial dosing requirement for warfarin. Many variants of uncertain importance were reported. INTERPRETATION: Although challenges remain, our results suggest that whole-genome sequencing can yield useful and clinically relevant information for individual patients. FUNDING: National Institute of General Medical Sciences; National Heart, Lung And Blood Institute; National Human Genome Research Institute; Howard Hughes Medical Institute; National Library of Medicine, Lucile Packard Foundation for Children's Health; Hewlett Packard Foundation; Breetwor Family Foundation.

Role of warfarin pharmacogenetic testing in clinical practice.

Chronic oral anticoagulation with warfarin is difficult to maintain within the therapeutic range and requires frequent monitoring and dose adjustments. Variations in two genes, VKORC1 and CYP2C9, have been associated with variation in warfarin metabolism among individuals. Patients with CYP2C9*2 and *3 variants have longer times to dose stabilization and are at higher risk of serious and life-threatening bleeding. VKORC1 polymorphisms significantly influence time to first therapeutic warfarin range, and variants in this gene determine low-, intermediate- and high-warfarin dose requirements. The prevalence of CYP2C9 and VKORC1 polymorphisms vary among different ethnic groups, and can account for over 30% of variance in warfarin dose. Recent studies suggest that the pharmacogenomics-guided dosing algorithm can accurately predict warfarin dosage and might reduce adverse events. We aim to review the pharmacogenetics of warfarin metabolism and the clinical role of genetic testing for warfarin therapy.

Comparison of assay systems for warfarin-related CYP2C9 and VKORC1 genotyping.

BACKGROUND: A variety of commercial genotyping assays is available to detect variants in the CYP2C9 and VKORC1 genes. The assay results are used in genotype-based warfarin dosing algorithms. We compared the performance of four such assay systems: Verigene, eSensor, Invader, and Luminex. METHODS: Result concordance and no call rates were determined on patient specimens tested on all four instruments. Turnaround times (TAT), hands-on time (HOT), pipetting steps and cost were obtained for runs of 1, 8 and 24 samples. RESULTS: The four assays were 100% concordant for the common CYP2C9 and VKORC1 alleles (n=100). Verigene had the shortest TAT and HOT for 1 and 8 samples. Verigene had the fewest pipetting steps for all sample sizes, while Invader had the most. Luminex had the longest TAT and highest cost for all sample run sizes. Verigene had the lowest cost for 1 and 8 samples and Invader the lowest for 24 samples. The no call rates for Verigene, Luminex, eSensor, and Invader were 10%, 4%, 1% and 0%, respectively. CONCLUSIONS: All assays gave comparable results for common variants. Each system offered unique advantages and disadvantages, whose relative importance depends on the needs of the adopting clinical laboratory.

Accuracy assessment of pharmacogenetically predictive warfarin dosing algorithms in patients of an academic medical center anticoagulation clinic.

The objectives of this retrospective cohort study are to evaluate the accuracy of pharmacogenetic warfarin dosing algorithms in predicting therapeutic dose and to determine if this degree of accuracy warrants the routine use of genotyping to prospectively dose patients newly started on warfarin. Seventy-one patients of an outpatient anticoagulation clinic at an academic medical center who were age 18 years or older on a stable, therapeutic warfarin dose with international normalized ratio (INR) goal between 2.0 and 3.0, and cytochrome P450 isoenzyme 2C9 (CYP2C9) and vitamin K epoxide reductase complex subunit 1 (VKORC1) genotypes available between January 1, 2007 and September 30, 2008 were included. Six pharmacogenetic warfarin dosing algorithms were identified from the medical literature. Additionally, a 5 mg fixed dose approach was evaluated. Three algorithms, Zhu et al. (Clin Chem 53:1199-1205, 2007), Gage et al. (J Clin Ther 84:326-331, 2008), and International Warfarin Pharmacogenetic Consortium (IWPC) (N Engl J Med 360:753-764, 2009) were similar in the primary accuracy endpoints with mean absolute error (MAE) ranging from 1.7 to 1.8 mg/day and coefficient of determination R (2) from 0.61 to 0.66. However, the Zhu et al. algorithm severely over-predicted dose (defined as >or=2x or >or=2 mg/day more than actual dose) in twice as many (14 vs. 7%) patients as Gage et al. 2008 and IWPC 2009. In conclusion, the algorithms published by Gage et al. 2008 and the IWPC 2009 were the two most accurate pharmacogenetically based equations available in the medical literature in predicting therapeutic warfarin dose in our study population. However, the degree of accuracy demonstrated does not support the routine use of genotyping to prospectively dose all patients newly started on warfarin.

A comparative study of most probable number (MPN)-PCR vs. real-time-PCR for the measurement of abundance and assessment of diversity of alkB homologous genes in soil.

The utilization of quantitative PCR (qPCR) approaches such as MPN-qPCR and real-time-qPCR for in situ assessment of functional genes yields substantial quantitative and qualitative differences. We show this by targeting the alkB gene related to biodegradation of aliphatic alkanes in a set of environmental samples with differing hydrocarbon content.

Assessment of deoxyhypusine hydroxylase as a putative, novel drug target.

Antimalarial drug resistance has nowadays reached each drug class on the market for longer than 10 years. The focus on validated, classical targets has severe drawbacks. If resistance is arising or already present in the field, a target-based High-Throughput-Screening (HTS) with the respective target involves the risk of identifying compounds to which field populations are also resistant. Thus, it appears that a rewarding albeit demanding challenge for target-based drug discovery is to identify novel drug targets. In the search for new targets for antimalarials, we have investigated the biosynthesis of hypusine, present in eukaryotic initiation factor 5A (eIF5A). Deoxyhypusine hydroxylase (DOHH), which has recently been cloned and expressed from P. falciparum, completes the modification of eIF5A through hydroxylation. Here, we assess the present druggable data on Plasmodium DOHH and its human counterpart. Plasmodium DOHH arose from a cyanobacterial phycobilin lyase by loss of function. It has a low FASTA score of 27 to its human counterpart. The HEAT-like repeats present in the parasite DOHH differ in number and amino acid identity from its human ortholog and might be of considerable interest for inhibitor design.