Estimation of monkeypox spread in a nonendemic country considering contact tracing and self-reporting: A stochastic modeling study.

In May 2022, monkeypox started to spread in nonendemic countries. To investigate contact tracing and self-reporting of the primary case in the local community, a stochastic model is developed. An algorithm based on Gillespie's stochastic chemical kinetics is used to quantify the number of infections, contacts, and duration from the arrival of the primary case to the detection of the index case (or until there are no more local infections). Different scenarios were set considering the delay in contact tracing and behavior of infectors. We found that the self-reporting behavior of a primary case is the most significant factor affecting outbreak size and duration. Scenarios with a self-reporting primary case have an 86% reduction in infections (average: 5-7, in a population of 10 000) and contacts (average: 27-72) compared with scenarios with a non-self-reporting primary case (average number of infections and contacts: 27-72 and 197-537, respectively). Doubling the number of close contacts per day is less impactful compared with the self-reporting behavior of the primary case as it could only increase the number of infections by 45%. Our study emphasizes the importance of the prompt detection of the primary case.

Multi-Faceted Analysis of COVID-19 Epidemic in Korea Considering Omicron Variant: Mathematical Modeling-Based Study.

BACKGROUND: The most recent variant of concern, omicron (B.1.1.529), has caused numerous cases worldwide including the Republic of Korea due to its fast transmission and reduced vaccine effectiveness. METHODS: A mathematical model considering age-structure, vaccine, antiviral drugs, and influx of the omicron variant was developed. We estimated transmission rates among age groups using maximum likelihood estimation for the age-structured model. The impact of non-pharmaceutical interventions (NPIs; in community and border), quantified by a parameter µ in the force of infection, and vaccination were examined through a multi-faceted analysis. A theory-based endemic equilibrium study was performed to find the manageable number of cases according to omicron- and healthcare-related factors. RESULTS: By fitting the model to the available data, the estimated values of µ ranged from 0.31 to 0.73, representing the intensity of NPIs such as social distancing level. If µ < 0.55 and 300,000 booster shots were administered daily from February 3, 2022, the number of severe cases was forecasted to exceed the severe bed capacity. Moreover, the number of daily cases is reduced as the timing of screening measures is delayed. If screening measure was intensified as early as November 24, 2021 and the number of overseas entrant cases was contained to 1 case per 10 days, simulations showed that the daily incidence by February 3, 2022 could have been reduced by 87%. Furthermore, we found that the incidence number in mid-December 2021 exceeded the theory-driven manageable number of daily cases. CONCLUSION: NPIs, vaccination, and antiviral drugs influence the spread of omicron and number of severe cases in the Republic of Korea. Intensive and early screening measures during the emergence of a new variant is key in controlling the epidemic size. Using the endemic equilibrium of the model, a formula for the manageable daily cases depending on the severity rate and average length of hospital stay was derived so that the number of severe cases does not surpass the severe bed capacity.

Risk estimation of the SARS-CoV-2 acute respiratory disease outbreak outside China.

BACKGROUND: On December 31, 2019, the World Health Organization was alerted to the occurrence of cases of pneumonia in Wuhan, Hubei Province, China, that were caused by an unknown virus, which was later identified as a coronavirus and named the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We aimed to estimate the reproductive number of SARS-CoV-2 in the Hubei Province and evaluate the risk of an acute respiratory coronavirus disease (COVID-19) outbreak outside China by using a mathematical model and stochastic simulations. RESULTS: We constructed a mathematical model of SARS-CoV-2 transmission dynamics, estimated the rate of transmission, and calculated the reproductive number in Hubei Province by using case-report data from January 11 to February 6, 2020. The possible number of secondary cases outside China was estimated by stochastic simulations in various scenarios of reductions in the duration to quarantine and rate of transmission. The rate of transmission was estimated as 0.8238 (95% confidence interval [CI] 0.8095-0.8382), and the basic reproductive number as 4.1192 (95% CI 4.0473-4.1912). Assuming the same rate of transmission as in Hubei Province, the possibility of no local transmission is 54.9% with a 24-h quarantine strategy, and the possibility of more than 20 local transmission cases is 7% outside of China. CONCLUSION: The reproductive number for SARS-CoV-2 transmission dynamics is significantly higher compared to that of the previous SARS epidemic in China. This implies that human-to-human transmission is a significant factor for contagion in Hubei Province. Results of the stochastic simulation emphasize the role of quarantine implementation, which is critical to prevent and control the SARS-CoV-2 outbreak outside China.

School Opening Delay Effect on Transmission Dynamics of Coronavirus Disease 2019 in Korea: Based on Mathematical Modeling and Simulation Study.

BACKGROUND: Nonpharmaceutical intervention strategy is significantly important to mitigate the coronavirus disease 2019 (COVID-19) spread. One of the interventions implemented by the government is a school closure. The Ministry of Education decided to postpone the school opening from March 2 to April 6 to minimize epidemic size. We aimed to quantify the school closure effect on the COVID-19 epidemic. METHODS: The potential effects of school opening were measured using a mathematical model considering two age groups: children (aged 19 years and younger) and adults (aged over 19). Based on susceptible-exposed-infectious-recovered model, isolation and behavior-changed susceptible individuals are additionally considered. The transmission parameters were estimated from the laboratory confirmed data reported by the Korea Centers for Disease Control and Prevention from February 16 to March 22. The model was extended with estimated parameters and estimated the expected number of confirmed cases as the transmission rate increased after school opening. RESULTS: Assuming the transmission rate between children group would be increasing 10 fold after the schools open, approximately additional 60 cases are expected to occur from March 2 to March 9, and approximately additional 100 children cases are expected from March 9 to March 23. After March 23, the number of expected cases for children is 28.4 for 7 days and 33.6 for 14 days. CONCLUSION: The simulation results show that the government could reduce at least 200 cases, with two announcements by the Ministry of education. After March 23, although the possibility of massive transmission in the children's age group is lower, group transmission is possible to occur.

Prioritization of vaccine strategy using an age-dependent mathematical model for 2009 A/H1N1 influenza in the Republic of Korea.

We developed a mathematical model of the 2009 A/H1N1 influenza epidemic in the Republic of Korea by considering five age groups and suggested the best way to prioritize an age-dependent vaccination strategy for mitigating the epidemic. An age-structured SEIAR influenza model was constructed based on the laboratory confirmed data obtained from the Korea Centers for Disease Control and Prevention (KCDC). The estimated transmission matrix captured one of the main characteristics of the 2009 A/H1N1 influenza, the transmission rate of which is high among young people, unlike that of seasonal influenza. We investigated the impact of age-dependent vaccination priority on the transmission dynamics of the 2009 A/H1N1 influenza and evaluated the Korean government vaccination policy when the vaccination started being administered 90 days (or 120 days) after the onset of the outbreak. We found that the government's age priority vaccination policy (Group 2, Group 1, Group 5, Group 4, and Group 3 in order) was a good strategy for reducing 62.06% of the cumulative cases when the vaccination was applied 90 days after the onset of the outbreak, while the proposed model's best suggestion (Group 2, Group 1, Group 3, Group 4, and Group 5 in order) showed 64.52% reduction. Furthermore, we studied the region-specific vaccination policy. For instance, the best age-priority of vaccination in Gwangwon province showed a different order (Group 3, Group 1, Group 2, Group 4, and Group 5 in order) and it reduced the incidence by 58.1%, which is 5.54% higher than that of the 2009 Korean government policy.

In vitro characterization of CYP102G4 from Streptomyces cattleya: A self-sufficient P450 naturally producing indigo.

Self-sufficient CYP102As possess outstanding hydroxylating activity to fatty acids such as myristic acid. Other CYP102 subfamily members share substrate specificity of CYP102As, but, occasionally, unusual characteristics of its own subfamily have been found. In this study, only one self-sufficient cytochrome P450 from Streptomyces cattleya was renamed from CYP102A_scat to CYP102G4, purified and characterized. UV-Vis spectrometry pattern, FAD/FMN analysis, and protein sequence comparison among CYP102s have shown that CYP102 from Streptomyces cattleya belongs to CYP102G subfamily. It showed hydroxylation activity toward fatty acids generating ω-1, ω-2, and ω-3-hydroxyfatty acids, which is similar to the general substrate specificity of CYP102 family. Unexpectedly, however, expression of CYP102G4 showed indigo production in LB medium batch flask culture, and high catalytic activity (kcat/Km) for indole was measured as 6.14±0.10min-1mM-1. Besides indole, CYP102G4 was able to hydroxylate aromatic compounds such as flavone, benzophenone, and chloroindoles. Homology model has shown such ability to accept aromatic compounds is due to its bigger active site cavity. Unlike other CYP102s, CYP102G4 did not have biased cofactor dependency, which was possibly determined by difference in NAD(P)H binding residues (Ala984, Val990, and Tyr1064) compared to CYP102A1 (Arg966, Lys972 and Trp1046). Overall, a self-sufficient CYP within CYP102G subfamily was characterized using purified enzymes, which appears to possess unique properties such as an only prokaryotic CYP naturally producing indigo.

Metapopulation model using commuting flow for national spread of the 2009 H1N1 influenza virus in the Republic of Korea.

We study the spatial-temporal pattern of the spread of the 2009 H1N1 influenza virus using a metapopulation model linked by commuting flow based on the reported influenza cases during the early stages of the epidemic in the Republic of Korea. The spatial heterogeneities, such as the local reproductive number and peak time, are investigated at province level. Furthermore, we discuss the effect of early intervention strategies, isolation and commuting restrictions, on the reduction of incidence at each province level. A major finding of this study is that early intervention at the source area of infection is more effective than interventions at the commuting-hub areas if the cost is limited.

Mathematical model and intervention strategies for mitigating tuberculosis in the Philippines.

Tuberculosis (TB) is the sixth leading cause of morbidity and mortality in the Philippines. Although significant progress has been made in the detection and cure of TB under the Directly Observed Treatment Short Course, battling against the disease is still a burdensome task. It demands a concerted effort for specific and effective interventions. In this work, a mathematical TB model fitted to the Philippine data is developed to understand its transmission dynamics. Different control strategies such as distancing, latent case finding, case holding, active case finding controls, and combinations thereof are investigated within the framework of optimal control theory. This study proposes optimal control strategies for reducing the number of high-risk latent and infectious TB patients with minimum intervention implementation costs. Results suggest that distancing control is the most efficient control strategy when a single intervention is utilized. However, full scale employment of the distancing control measure is a daunting task. This burden can be circumvented by the combination of other control interventions. Our noble finding in this study is that enhancing active case finding control instead of case holding control together with distancing and latent case finding control is shown to have significant potential for curtailing the spread of TB in the Philippines.

Semi-rational engineering of CYP153A35 to enhance ω-hydroxylation activity toward palmitic acid.

CYP153A35 from Gordonia alkanivorans was recently characterized as fatty acid ω-hydroxylase. To enhance the catalytic activity of CYP153A35 toward palmitic acid, site-directed saturation mutagenesis was attempted using a semi-rational approach that combined structure-based computational analysis and subsequent saturation mutagenesis. Using colorimetric high-throughput screening (HTS) method based on O-demethylation activity of P450, CYP153A35 D131S and D131F mutants were selected. The best mutant, D131S, having a single mutation on BC-loop, showed 13- and 17-fold improvement in total turnover number (TTN) and catalytic efficiency (k cat/K M) toward palmitic acid compared to wild-type, respectively. However, in whole-cell reaction, D131S mutant showed only 50% improvement in ω-hydroxylated palmitic acid yield compared to the wild type. Docking simulation studies explained that the effect of D131S mutation on the catalytic activity would be mainly caused by the binding pose of fatty acids in the substrate access tunnel of the enzyme. This effect of D131S mutation on the catalytic activity is synergistic with that of the mutations in the active site previously reported.

Mathematical model of transmission dynamics and optimal control strategies for 2009 A/H1N1 influenza in the Republic of Korea.

A mathematical model for the transmission dynamics of the 2009 A/H1N1 influenza epidemic in the Republic of Korea is developed. The simulation period is separated into three consecutive periods based on the government's intervention strategies: the nonpharmaceutical strategy is used during Period 1. The nonpharmaceutical and antiviral strategies are executed during Period 2 and the vaccine strategy is added during Period 3. During Period 1, we estimate the reduction in the transmission rate due to the government's intervention policies as a difference between the data-fitted and uncontrolled transmission rate that is derived from the basic reproductive number, R0, of the model without intervention. This quantified reduced transmission rate is used as an upperbound of the nonpharmaceutical control for studying optimal control strategies, which is a new approach for determining the realistic upperbound of control. In this study, we also explore the real-time prediction of incidence using the mathematical model during the early stage of the epidemic. We investigate the impact of vaccination coverage and timing with respect to the cumulative incidence. The result implies that early vaccination plays a significant role for preventing the epidemic.

P212A Mutant of Dihydrodaidzein Reductase Enhances (S)-Equol Production and Enantioselectivity in a Recombinant Escherichia coli Whole-Cell Reaction System.

(S)-Equol, a gut bacterial isoflavone derivative, has drawn great attention because of its potent use for relieving female postmenopausal symptoms and preventing prostate cancer. Previous studies have reported on the dietary isoflavone metabolism of several human gut bacteria and the involved enzymes for conversion of daidzein to (S)-equol. However, the anaerobic growth conditions required by the gut bacteria and the low productivity and yield of (S)-equol limit its efficient production using only natural gut bacteria. In this study, the low (S)-equol biosynthesis of gut microorganisms was overcome by cloning the four enzymes involved in the biosynthesis from Slackia isoflavoniconvertens into Escherichia coli BL21(DE3). The reaction conditions were optimized for (S)-equol production from the recombinant strain, and this recombinant system enabled the efficient conversion of 200 µM and 1 mM daidzein to (S)-equol under aerobic conditions, achieving yields of 95% and 85%, respectively. Since the biosynthesis of trans-tetrahydrodaidzein was found to be a rate-determining step for (S)-equol production, dihydrodaidzein reductase (DHDR) was subjected to rational site-directed mutagenesis. The introduction of the DHDR P212A mutation increased the (S)-equol productivity from 59.0 mg/liter/h to 69.8 mg/liter/h in the whole-cell reaction. The P212A mutation caused an increase in the (S)-dihydrodaidzein enantioselectivity by decreasing the overall activity of DHDR, resulting in undetectable activity for (R)-dihydrodaidzein, such that a combination of the DHDR P212A mutant with dihydrodaidzein racemase enabled the production of (3S,4R)-tetrahydrodaidzein with an enantioselectivity of >99%.

Production of p-hydroxybenzoic acid from p-coumaric acid by Burkholderia glumae BGR1.

p-Coumaric acid (pCA) is abundant in biomass with low lignin content, such as straw and stubble from rye, wheat, and barley. pCA can be isolated from biomass and used for the synthesis of aromatic hydrocarbons. Here, we report engineering of the natural pathway for conversion of pCA into p-hydroxybenzoic acid (pHBA) to increase the amount of pHBA that accumulates more than 100-fold. Burkholderia glumae strain BGR1 (BGR1) grows efficiently on pCA as a sole carbon source via a CoA-dependent non-ß-oxidation pathway. This pathway removes two carbons from pCA as acetyl-CoA yielding p-hydroxybenzaldehyde and subsequently oxidizes it to pHBA. To increase the amount of accumulated pHBA in BGR1, we first deleted two genes encoding enzymes that degrade pHBA in the ß-ketoadipate pathway. At 10 mM of pCA, the double deletion mutant BGR1_PB4 (Δphb3hΔbcl) accumulated pHBA with 95% conversion, while the control BGR1 accumulated only with 11.2% conversion. When a packed bed reactor containing immobilized BGR1_PB4 cells was operated at a dilution rate 0.2 h(-1) , the productivity of pHBA was achieved at 9.27 mg/L/h for 134 h. However, in a batch reactor at 20 mM pCA, growth of BGR1_PB4 was strongly inhibited, resulting in a low conversion of 19.3%. To further increase the amount of accumulated pCA, we identified the first enzyme in the pathway, p-hydroxcinnmaoyl-CoA synthetase II (phcs II), as the rate-limiting enzyme. Over expression of phcs II using a Palk promoter in a batch reaction at 20 mM of pCA yielded 99.0% conversion to pHBA, which is the highest concentration of pHBA ever reported using a biological process. Biotechnol. Bioeng. 2016;113: 1493-1503. © 2015 Wiley Periodicals, Inc.

A dynamic compartmental model for the Middle East respiratory syndrome outbreak in the Republic of Korea: A retrospective analysis on control interventions and superspreading events.

The 2015 Middle East respiratory syndrome (MERS) outbreak in the Republic of Korea has provided an opportunity to improve our understanding of the spread of MERS linked to healthcare settings. Here we designed a dynamic transmission model to analyze the MERS outbreak in the Republic of Korea based on confirmed cases reported during the period May 20-July 4, 2015. Our model explicitly incorporates superspreading events and time-dependent transmission and isolation rates. Our model was able to provide a good fit to the trajectory of the outbreak and was useful to analyze the role of hypothetical control scenarios. Specifically, we assessed the impact of the timing of control measures, especially associated with a reduction of the transmission rate and diagnostic delays on outbreak size and duration. Early interventions within 1week after the epidemic onset, for instance, including the initial government announcement to the public about the list of hospitals exposed to MERS coronavirus (MERS-CoV), show a promising means to reduce the size (>71%) and duration (>35%) of the MERS epidemic. Finally, we also present results of an uncertainty analysis focused on the role of superspreading events.

Production of ω-hydroxy palmitic acid using CYP153A35 and comparison of cytochrome P450 electron transfer system in vivo.

Bacterial cytochrome P450 enzymes in cytochrome P450 (CYP)153 family were recently reported as fatty acid ω-hydroxylase. Among them, CYP153As from Marinobacter aquaeolei VT8 (CYP153A33), Alcanivorax borkumensis SK2 (CYP153A13), and Gordonia alkanivorans (CYP153A35) were selected, and their specific activities and product yields of ω-hydroxy palmitic acid based on whole cell reactions toward palmitic acid were compared. Using CamAB as redox partner, CYP153A35 and CYP153A13 showed the highest product yields of ω-hydroxy palmitic acid in whole cell and in vitro reactions, respectively. Artificial self-sufficient CYP153A35-BMR was constructed by fusing it to the reductase domain of CYP102A1 (i.e., BM3) from Bacillus megaterium, and its catalytic activity was compared with CYP153A35 and CamAB systems. Unexpectedly, the system with CamAB resulted in a 1.5-fold higher yield of ω-hydroxy palmitic acid than that using A35-BMR in whole cell reactions, whereas the electron coupling efficiency of CYP153A35-BM3 reductase was 4-fold higher than that of CYP153A35 and CamAB system. Furthermore, various CamAB expression systems according to gene arrangements of the three proteins and promoter strength in their gene expression were compared in terms of product yields and productivities. Tricistronic expression of the three proteins in the order of putidaredoxin (CamB), CYP153A35, and putidaredoxin reductase (CamA), i.e., A35-AB2, showed the highest product yield from 5 mM palmitic acid for 9 h in batch reaction owing to the concentration of CamB, which is the rate-limiting factor for the activity of CYP153A35. However, in fed-batch reaction, A35-AB1, which expressed the three proteins individually using three T7 promoters, resulted with the highest product yield of 17.0 mM (4.6 g/L) ω-hydroxy palmitic acid from 20 mM (5.1 g/L) palmitic acid for 30 h.

Transmission characteristics of MERS and SARS in the healthcare setting: a comparative study.

BACKGROUND: The Middle East respiratory syndrome (MERS) coronavirus has caused recurrent outbreaks in the Arabian Peninsula since 2012. Although MERS has low overall human-to-human transmission potential, there is occasional amplification in the healthcare setting, a pattern reminiscent of the dynamics of the severe acute respiratory syndrome (SARS) outbreaks in 2003. Here we provide a head-to-head comparison of exposure patterns and transmission dynamics of large hospital clusters of MERS and SARS, including the most recent South Korean outbreak of MERS in 2015. METHODS: To assess the unexpected nature of the recent South Korean nosocomial outbreak of MERS and estimate the probability of future large hospital clusters, we compared exposure and transmission patterns for previously reported hospital clusters of MERS and SARS, based on individual-level data and transmission tree information. We carried out simulations of nosocomial outbreaks of MERS and SARS using branching process models rooted in transmission tree data, and inferred the probability and characteristics of large outbreaks. RESULTS: A significant fraction of MERS cases were linked to the healthcare setting, ranging from 43.5 % for the nosocomial outbreak in Jeddah, Saudi Arabia, in 2014 to 100 % for both the outbreak in Al-Hasa, Saudi Arabia, in 2013 and the outbreak in South Korea in 2015. Both MERS and SARS nosocomial outbreaks are characterized by early nosocomial super-spreading events, with the reproduction number dropping below 1 within three to five disease generations. There was a systematic difference in the exposure patterns of MERS and SARS: a majority of MERS cases occurred among patients who sought care in the same facilities as the index case, whereas there was a greater concentration of SARS cases among healthcare workers throughout the outbreak. Exposure patterns differed slightly by disease generation, however, especially for SARS. Moreover, the distributions of secondary cases per single primary case varied highly across individual hospital outbreaks (Kruskal-Wallis test; P < 0.0001), with significantly higher transmission heterogeneity in the distribution of secondary cases for MERS than SARS. Simulations indicate a 2-fold higher probability of occurrence of large outbreaks (>100 cases) for SARS than MERS (2 % versus 1 %); however, owing to higher transmission heterogeneity, the largest outbreaks of MERS are characterized by sharper incidence peaks. The probability of occurrence of MERS outbreaks larger than the South Korean cluster (n = 186) is of the order of 1 %. CONCLUSIONS: Our study suggests that the South Korean outbreak followed a similar progression to previously described hospital clusters involving coronaviruses, with early super-spreading events generating a disproportionately large number of secondary infections, and the transmission potential diminishing greatly in subsequent generations. Differences in relative exposure patterns and transmission heterogeneity of MERS and SARS could point to changes in hospital practices since 2003 or differences in transmission mechanisms of these coronaviruses.

A spatial-temporal transmission model and early intervention policies of 2009 A/H1N1 influenza in South Korea.

We developed a spatial-temporal model of the 2009 A/H1N1 influenza pandemic in the Seoul metropolitan area (SMA), which is located in the north-west of South Korea and is the second-most complex metropolitan area worldwide. This multi-patch influenza model consists of a SEIAR influenza transmission model and flow model between two districts. This model is based on the daily confirmed cases of A/H1N1 influenza collected by the Korea Center for Disease Control and Prevention from April 27 to September 15, 2009 and the daily commuting data from 33 districts of SMA reported in the 2010 Population and Housing Census (PHC). We analyzed the spread patterns of 2009 influenza in the SMA by the reproductive numbers and geographic information systems. During the early period of novel influenza pandemics, when pharmaceutical interventions are lacking, non-pharmaceutical public health interventions will be the most critical strategies for impeding the spread of influenza and delaying an epidemic. Using the spatial-temporal model developed herein, we also investigated the impact of non-pharmaceutical public health interventions, isolation and/or commuting restrictions, on the incidence reduction in various scenarios. Our model provides scientific evidence for predicting the spread of disease and preparedness for a future pandemic.

Optimal intervention strategy for prevention tuberculosis using a smoking-tuberculosis model.

In this paper, we developed a dynamic model of smoking-tuberculosis (TB) transmission in South Korea, and investigated the effects of control strategies on the number of incidence of TB using optimal control theory. Model parameters regarding TB and smoking are estimated through least-squares fitting to real data. We considered three TB controls (distancing, case-finding, and case-holding) and two smoking controls (distancing and quitting), in order to minimize the number of exposed and infectious individuals and the cost of control. Numerical simulations for the various control strategies highlight that implementing the smoking controls, not with TB controls, can effectively reduce the incidence of TB transmission.

Fungal cytochrome P450 monooxygenases of Fusarium oxysporum for the synthesis of ω-hydroxy fatty acids in engineered Saccharomyces cerevisiae.

BACKGROUND: Omega hydroxy fatty acids (ω-OHFAs) are multifunctional compounds that act as the basis for the production of various industrial products with broad commercial and pharmaceutical implications. However, the terminal oxygenation of saturated or unsaturated fatty acids for the synthesis of ω-OHFAs is intricate to accomplish through chemocatalysis, due to the selectivity and controlled reactivity in C-H oxygenation reactions. Cytochrome P450, the ubiquitous enzyme is capable of catalyzing the selective terminal omega hydroxylation naturally in biological kingdom. RESULTS: To gain a deep insight on the biochemical role of fungal P450s towards the production of omega hydroxy fatty acids, two cytochrome P450 monooxygenases from Fusarium oxysporum (FoCYP), FoCYP539A7 and FoCYP655C2; were identified, cloned, and heterologously expressed in Saccharomyces cerevisiae. For the efficient production of ω-OHFAs, the S. cerevisiae was engineered to disrupt the acyl-CoA oxidase enzyme and the ß-oxidation pathway inactivated (ΔPox1) S. cerevisiae mutant was generated. To elucidate the significance of the interaction of redox mechanism, FoCYPs were reconstituted with the heterologous and homologous reductase systems--S. cerevisiae CPR (ScCPR) and F. oxysporum CPR (FoCPR). To further improve the yield, the effect of pH was analyzed and the homologous FoCYP-FoCPR system efficiently hydroxylated caprylic acid, capric acid and lauric acid into their respective ω-hydroxy fatty acids with 56%, 79% and 67% conversion. Furthermore, based on computational simulations, we identified the key residues (Asn106 of FoCYP539A7 and Arg235 of FoCYP655C2) responsible for the recognition of fatty acids and demonstrated the structural insights of the active site of FoCYPs. CONCLUSION: Fungal CYP monooxygenases, FoCYP539A7 and FoCYP655C2 with its homologous redox partner, FoCPR constitutes a promising catalyst due to its high regio- and stereo-selectivity in the hydroxylation of fatty acids and in the substantial production of industrially valuable ω-hydroxy fatty acids.

A Multiscale Model of Cardiovascular System Including an Immersed Whole Heart in the Cases of Normal and Ventricular Septal Defect (VSD).

A mathematical and computational model combining the heart and circulatory system has been developed to understand the hemodynamics of circulation under normal conditions and ventricular septal defect (VSD). The immersed boundary method has been introduced to describe the interaction between the moving two-dimensional heart and intracardiac blood flow. The whole-heart model is governed by the Navier-Stokes system; this system is combined with a multi-compartment model of circulation using pressure-flow relations and the linearity of the discretized Navier-Stokes system. We investigate the velocity field, flowmeters, and pressure-volume loop in both normal and VSD cases. Simulation results show qualitatively good agreements with others found in the literature. This model, combining the heart and circulation, is useful for understanding the complex, hemodynamic mechanisms involved in normal circulation and cardiac diseases.

The production of ω-hydroxy palmitic acid using fatty acid metabolism and cofactor optimization in Escherichia coli.

Hydroxylated fatty acids (HFAs) are used as important precursors for bulk and fine chemicals in the chemical industry. Here, to overproduce long-chain (C16-C18) fatty acids and hydroxy fatty acid, their biosynthetic pathways including thioesterase (Lreu_0335) from Lactobacillus reuteri DSM20016, ß-hydroxyacyl-ACP dehydratase (fabZ) from Escherichia coli, and a P450 system (i.e., CYP153A from Marinobacter aquaeolei VT8 and camA/camB from Pseudomonas putida ATCC17453) were overexpressed. Acyl-CoA synthase (fadD) involved in fatty acid degradation by ß-oxidation was also deleted in E. coli BW25113. The engineered E. coli FFA4 strain without the P450 system could produce 503.0 mg/l of palmitic (C16) and 508.4 mg/l of stearic (C18) acids, of which the amounts are ca. 1.6- and 2.3-fold higher than those of the wild type. On the other hand, the E. coli HFA4 strain including the P450 system for ω-hydroxylation could produce 211.7 mg/l of ω-hydroxy palmitic acid, which was 42.1 ± 0.1 % of the generated palmitic acid, indicating that the hydroxylation reaction was the rate-determining step for the HFA production. For the maximum production of ω-hydroxy palmitic acid, NADH, i.e., an essential cofactor for P450 reaction, was overproduced by the integration of NAD(+)-dependent formate dehydrogenase (FDH) from Candida boidinii into E. coli chromosome and the deletion of alcohol dehydrogenase (ADH). Finally, the NADH-level-optimized E. coli strain produced 610 mg/l of ω-hydroxy palmitic acid (ω-HPA), which was almost a threefold increase in its yield compared to the same strain without NADH overproduction.