Filamin A forms a complex with EphA2 and regulates EphA2 serine 897 phosphorylation and glioblastoma cell proliferation.

EphA2 is phosphorylated on serine 897 (S897) in response to growth factors such as epidermal growth factor (EGF) and on tyrosine 588 (Y588) in response to its ligand ephrinA1, causing different cellular responses. In this study, we show that the actin-binding protein Filamin A forms a complex with EphA2 and promotes its S897 phosphorylation and glioblastoma cell proliferation. Suppression of Filamin A expression by siRNAs inhibited glioblastoma cell proliferation induced by EGF stimulation or overexpression of EphA2. Knockdown of Filamin A inhibited EGF-induced S897 phosphorylation of EphA2, whereas it had little effect on ephrinA1-induced Y588 phosphorylation of EphA2. Furthermore, Filamin A expression affected the subcellular localization of EphA2. This study suggests that Filamin A selectively promotes EphA2 S897 phosphorylation and plays an important role in glioblastoma cell proliferation.

Low Serum Zinc Concentration Is Associated With Infection Particularly in Patients With Stage 5 Chronic Kidney Disease Medicated with Proton Pump Inhibitors.

OBJECTIVE: Zinc (Zn) plays an important role in immune function. Several studies have identified an association between a Zn deficiency and infection. Infectious diseases are major complications of chronic kidney disease (CKD). We investigated whether serum Zn concentrations are associated with risk of infection in patients with advanced CKD. DESIGN AND METHODS: We retrospectively analyzed data from 299 patients with CKD whose serum Zn values were measured to evaluate anemia between January 2013 and December 2016. Among them, 9 who were supplemented with Zn and 67 who had started urgent dialysis at the time of measurement were excluded. We analyzed infection events, length of infection-related hospitalization and infection-related and all-cause mortality in the remaining 223 patients during a median follow-up of 36 months. We assigned the patients to groups with low or high Zn values (≤50 and >50 µg/dL, respectively) based on a median value of 50 µg/dL. Data were analyzed using Kaplan-Meier curves and Cox hazards models. RESULTS: During a median follow-up of 36 months, 40 patients were hospitalized with infections. The rate of infection-related and long-term hospitalization (>10 days) due to infection was higher for patients with low, than high, Zn values (23.3% vs. 12.6%; P = .042 and 26.2% vs. 12.4%; P = .007, respectively). After adjustment in Cox hazards models, low serum Zn values remained an independent risk factor for infection-related hospitalization (Hazard ratio [HR], 1.93; 95% confidence interval [CI], 1.01-3.71; P = .048), especially for patients on proton pump inhibitor (PPI) medications (HR, 2.66, 95%; CI, 1.22-5.81; P = .014). CONCLUSION: Patients with advanced CKD accompanied by low serum Zn concentration, and particularly those medicated with PPI, are at high risk of infection-related hospitalization, which results in long-term hospitalization.

A novel risk score on admission for predicting death or need for surgery in patients with acute type A intramural hematoma receiving medical therapy.

There has been continuing discussion regarding the treatment strategy for acute type A intramural hematoma (IMH). Most patients are treated conservatively in Japan; hence, predicting fatal events and stratifying risks based on results normally obtained on hospital arrival are important. We aimed to examine the incidences and risk factors of death or need for surgery for acute type A IMH in patients receiving medical treatment and to identify high-risk patients using clinical findings on hospital arrival. From 2011 to 2016, 57 consecutive patients (mean age 72.5 years; male 36.8%) diagnosed with acute type A IMH who were receiving treatment at Shizuoka City Shizuoka Hospital were retrospectively included. Primary endpoint was a composite of cardiovascular death and operation within 1 year after onset. To evaluate sensitivity and specificity of the risk factors and risk score, we estimated the area under the receiver operating characteristic (ROC) curve. Mean follow-up duration was 621 days. Mean systolic blood pressure (SBP) was 129 mmHg. Computed tomography (CT) on arrival showed a mean ascending aorta diameter of 46 mm. Ulcer-like projection (ULP) in the ascending aorta and pericardial effusion (PE) were seen in 33% and 42% of cases, respectively. Twenty-eight patients (49.1%) reached the primary endpoint (cardiovascular death, 7 cases [12.3%]; operation, 21 cases [36.8%]). In univariate analysis of admission values, the primary endpoint group had significantly lower SBP (113.0 ± 28.5 vs 144.3 ± 33.5 mmHg), higher ascending aorta diameter (49.5 ± 8.1 vs 43.6 ± 5.9 mm), and higher frequency of ULP (53.8% vs 13.8%) and PE (58.6% vs 25.0%) than the event-free group. Multivariate analysis showed that ULP on admission CT was a significant predictor of the primary endpoint. The risk score was considered using these risk factors. On admission, the primary endpoint could be predicted with 89.7% sensitivity and 75% specificity (area under the ROC curve 0.823) if the patient had ULP and/or > 2 of the following factors: SBP < 120 mmHg, ascending aorta diameter > 45 mm, and PE. SBP and CT findings on arrival were significantly associated with cardiovascular death and the need for surgery in patients with acute type A IMH receiving initial medical therapy. The novel risk score was useful for predicting cardiovascular death and surgery.

Quality of life of elderly patients on peritoneal dialysis versus hemodialysis: a single-center study.

BACKGROUND: The number of elderly dialysis patients in Japan is dramatically increasing. Receiving therapy with better satisfaction through home care is one of the important factors in their daily lives. Thus, the quality of life of elderly patients on hemodialysis (HD) or peritoneal dialysis (PD) was evaluated. METHODS: Clinical information of patients aged ≥80 years who started dialysis at our hospital between January 2013 and December 2015 was retrospectively collected. The mortality rate, length of hospitalization, and place of death were identified to evaluate patient quality of life. RESULTS: In total, 56 patients (14 PD and 42 HD) were enrolled. The mean age of study subjects was 85.2 ± 4.0 years. The proportion of PD patients who lived with their family or have professional caregivers who could assist them in their daily life was higher than that of HD patients (100 vs. 76.2%, respectively; p = 0.044). Mortality rate was higher in PD patients than in HD patients (p = 0.003), but long-term hospitalization of >180 days was observed only in HD patients (PD vs. HD: 0.0 vs. 16.7%; p = 0.102). In patients with Barthel index scores <100, the long-term hospitalization difference was significant (PD vs. HD: 0.0 vs. 30.4%; p = 0.040). Of note, 6 of 7 deceased PD patients and 1 of 10 deceased HD patients died at home (p = 0.002). CONCLUSION: PD is a desirable home care therapy for elderly patients, but the burden on caregivers should be considered.

A prospective observational study of early intervention with erythropoietin therapy and renal survival in non-dialysis chronic kidney disease patients with anemia: JET-STREAM Study.

BACKGROUND: There is limited data showing that early treatment for anemia could prolong renal survival in non-dialysis chronic kidney disease (CKD) patients. We therefore investigated the relationship between hemoglobin (Hb) levels at initiation of epoetin beta therapy and renal outcome in non-dialysis CKD patients with anemia. METHODS: In this prospective, multi-center, observational study, non-dialysis CKD patients with anemia who were naïve to erythropoiesis-stimulating agents (ESAs) were divided into three groups based on their Hb levels at initiation of epoetin beta therapy (Group I: 10 ≤ Hb < 11 g/dL, Group II: 9 ≤ Hb < 10 g/dL, and Group III: Hb < 9 g/dL). The primary endpoint was time to first occurrence of any renal event. For the primary analysis, an inverse probability weighted Cox regression model was used to adjust time-dependent selection bias in the artificially censored data. RESULTS: A total of 1113 patients were eligible for primary endpoint analysis. Risk of renal events was significantly higher in Group III compared with Group I (HR, 2.52; 95 % CI, 1.98-3.21; P < 0.0001); although not significant, the risk was also higher in Group II compared with Group I (HR, 1.48; 95 % CI, 0.91-2.40; P = 0.11). CONCLUSION: Initiation of ESA therapy when Hb levels decreased below 11 g/dL but not below 10 g/dL could be more effective at reducing the risk of renal events in non-dialysis CKD patients with anemia compared with initiation of ESA therapy at below 9 g/dL or even 10 g/dL.

X-linked RNA-binding motif protein (RBMX) is required for the maintenance of Borna disease virus nuclear viral factories.

Borna disease virus (BDV) is a non-segmented, negative-strand RNA virus that establishes persistent infection in the nucleus. Although BDV forms viral inclusion bodies, termed viral speckles of transcripts (vSPOTs), which are associated with chromatin in the nucleus, the host factors involved in the maintenance of vSPOTs remain largely unknown. In this study, we identified X-linked RNA-binding motif protein (RBMX) as a nuclear factor interacting with BDV nucleoprotein. Interestingly, knockdown of RBMX led to disruption of the formation of vSPOTs and reduced both transcription and replication of BDV. Our results indicate that RBMX is involved in the maintenance of the structure of the virus factory in the nucleus.

Dialysis fluid endotoxin level and mortality in maintenance hemodialysis: a nationwide cohort study.

BACKGROUND: The quality of dialysis fluid water might play an important role in hemodialysis patient outcomes. Although targeted endotoxin levels of dialysis fluid vary among countries, evidence of the contribution of these levels to mortality in hemodialysis patients is lacking. STUDY DESIGN: Retrospective cohort study using data from the Japan Renal Data Registry, a nationwide annual survey. SETTING & PARTICIPANTS: 130,781 patients receiving thrice-weekly in-center hemodialysis for more than 6 months were enrolled at 2,746 facilities in Japan at the end of 2006. None of the patients changed facility or treatment modality during 2007. PREDICTOR: Highest endotoxin level in dialysis fluid reported by each facility during 2006. Patients were categorized by facility endotoxin level into the following groups: <0.001, 0.001 to <0.01, 0.01 to <0.05, 0.05 to <0.1, and ≥0.1EU/mL. Age, sex, dialysis vintage, diabetes mellitus as a primary cause of end-stage renal disease, Kt/V, normalized protein catabolic rate, dialysis session duration, serum albumin, and hemoglobin were measured as potential confounders. OUTCOME: All-cause mortality, censored by transplantation; withdrawal from dialysis treatment; or end of follow-up. RESULTS: Of 130,781 hemodialysis patients, 91.2% had facility endotoxin levels below the limit set for dialysis fluid in Japan (<0.05EU/mL). During a 1-year follow-up, 8,978 (6.9%) patients died of all causes. The rate of all-cause mortality at 1 year was highest in the ≥0.1-EU/mL category (88.0 deaths/1,000 person-years). Patients in the ≥0.1-EU/mL group exhibited an increased risk of all-cause mortality of 28% (95% CI, 10%-48%) compared to the <0.001-EU/mL group. LIMITATIONS: Endotoxin level in dialysis fluid is reported as categorical data. No information about variation in endotoxin levels in dialysis fluid over time. CONCLUSIONS: Higher facility endotoxin levels in dialysis fluid may be related to increased risk for all-cause mortality among hemodialysis patients. Correcting this modifiable facility water management practice might improve the outcome of hemodialysis patients.

Nucleolar scaffold protein, WDR46, determines the granular compartmental localization of nucleolin and DDX21.

The nuclear scaffold is an insoluble nuclear structure that contributes to the inner nuclear organization. In this study, we showed that one of the nuclear scaffold proteins, WDR46, plays a role as a fundamental scaffold component of the nucleolar structure. WDR46 is a highly insoluble nucleolar protein, and its subcellular localization is dependent on neither DNA nor RNA. The N- and C-terminal regions of WDR46 are predicted to be intrinsically disordered, and both regions are critical for the nucleolar localization of WDR46 and the association with its binding partners. When WDR46 was knocked down, two of its binding partners, nucleolin and DDX21 (involved in 18S rRNA processing), were mislocalized from the granular component to the edges of the nucleoli, whereas other binding partners, NOP2 and EBP2 (involved in 28S rRNA processing), were not affected. This is because the proper recruitment of nucleolin and DDX21 to the nucleoli in daughter cells after cell division is ensured by WDR46. These findings suggest a structural role for WDR46 in organizing the 18S ribosomal RNA processing machinery. This role of WDR46 is enabled by its interaction property via intrinsically disordered regions.

Ten-Gram-Scale Total Synthesis of the Anticancer Drug Candidate E7130 to Supply Clinical Trials.

E7130 is a novel drug candidate with an exceedingly complex chemical structure of the halichondrin class, discovered by a total synthesis approach through joint research between the Kishi group at Harvard University and Eisai. Only 18 months after completion of the initial milligram-scale synthesis, ten-gram-scale synthesis of E7130 was achieved, providing the first good manufacturing practice (GMP) batch to supply clinical trials. This paper highlights the challenges in developing ten-gram-scale synthesis from the milligram-scale synthesis.

The interferon stimulated gene-encoded protein HELZ2 inhibits human LINE-1 retrotransposition and LINE-1 RNA-mediated type I interferon induction.

Some interferon stimulated genes (ISGs) encode proteins that inhibit LINE-1 (L1) retrotransposition. Here, we use immunoprecipitation followed by liquid chromatography-tandem mass spectrometry to identify proteins that associate with the L1 ORF1-encoded protein (ORF1p) in ribonucleoprotein particles. Three ISG proteins that interact with ORF1p inhibit retrotransposition: HECT and RLD domain containing E3 ubiquitin-protein ligase 5 (HERC5); 2'-5'-oligoadenylate synthetase-like (OASL); and helicase with zinc finger 2 (HELZ2). HERC5 destabilizes ORF1p, but does not affect its cellular localization. OASL impairs ORF1p cytoplasmic foci formation. HELZ2 recognizes sequences and/or structures within the L1 5'UTR to reduce L1 RNA, ORF1p, and ORF1p cytoplasmic foci levels. Overexpression of WT or reverse transcriptase-deficient L1s lead to a modest induction of IFN-α expression, which is abrogated upon HELZ2 overexpression. Notably, IFN-α expression is enhanced upon overexpression of an ORF1p RNA binding mutant, suggesting ORF1p binding might protect L1 RNA from "triggering" IFN-α induction. Thus, ISG proteins can inhibit retrotransposition by different mechanisms.

Long-Term Proton Pump Inhibitor Therapy Increases the Risk of Infection in Patients with Incident Hemodialysis.

BACKGROUND: Infection is one of the most common causes of death in patients with chronic kidney disease (CKD). Proton pump inhibitors (PPIs) are not only widely used in patients with CKD but also represent a known risk factor for infection in the general population. Here, we investigated associations between PPIs and infection events in patients with incident hemodialysis. METHODS: We analyzed data from 485 consecutive patients with CKD who started hemodialysis at our hospital between January 2013 and December 2019. We analyzed associations between infection events and long-term (≥6 months) PPI use before and after propensity score-matched analysis. RESULTS: Of the 485 patients, PPIs were administered to 177 patients (36.5%). During 24 months of follow-up, infection events occurred in 53 patients (29.9%) with PPIs and 40 patients (13.0%) without PPIs (p < 0.001). Patients with PPIs had a significantly higher cumulative incidence rate of infection events than those without PPIs (hazard ratio [HR] 2.13, 95% confidence interval [CI]: 1.36-3.32; p < 0.001). Even after propensity score-matched analysis (132 patients matched in each), the rate of infection events was higher for patients with PPIs (28.8% vs. 12.1%, HR 2.88, 95% CI: 1.61-5.16; p < 0.001). Similar results were obtained for severe infection events in both unmatched (14.1% vs. 4.5%, HR 2.97, 95% CI: 1.47-6.00; p = 0.002) and propensity score-matched analyses (14.4% vs. 3.8%, HR 4.54, 95% CI: 1.85-11.13; p < 0.001). CONCLUSIONS: In patients with incident hemodialysis, long-term PPI use increases the risk of infection. Clinicians should be wary of unnecessarily prolonging PPI therapy.

Structure of N-formylglycinamide ribonucleotide amidotransferase II (PurL) from Thermus thermophilus HB8.

The crystal structure of PurL from Thermus thermophilus HB8 (TtPurL; TTHA1519) was determined in complex with an adenine nucleotide, PO(4)(3-) and Mg(2+) at 2.35 Å resolution. TtPurL consists of 29 α-helices and 28 ß-strands, and one loop is disordered. TtPurL consists of four domains, A1, A2, B1 and B2, and the structures of the A1-B1 and A2-B2 domains were almost identical to each other. Although the sequence identity between TtPurL and PurL from Thermotoga maritima (TmPurL) is higher than that between TtPurL and the PurL domain of the large PurL from Salmonella typhimurium (StPurL), the secondary structure of TtPurL is much more similar to that of StPurL than to that of TmPurL.

Bacteriological water quality in the central dialysis fluid delivery system from the survey of the Japanese Society for Dialysis Therapy.

The Japanese Society for Dialysis Therapy (JSDT) surveyed all dialysis facilities for bacteriological quality of dialysis fluid and quality controls for dialysis fluid in 2006 and 2007. The JSDT collected the data for endotoxin (ET) levels, bacterial count and usage of ET retentive filters (ETRF). The JSDT standard for ET level in dialysis fluid (<0.050 EU/ml) was achieved in 89.0% in 2006 and in 93.6% in 2007. The JSDT standard for bacterial cell counts in dialysis fluid (<100 cfu/ml) was achieved in 96.9% in 2006 and in 97.4% in 2007. The central dialysis fluid delivery system (CDDS) is a unique system developed in Japan which has easy handling for daily maintenance of delivery systems, but it has been pointed out that CDDS has a weak point for the protection of biofilms. However, the bacteriological water qualities of dialysis fluid in CDDS were proven to be extremely high in most Japanese dialysis facilities by JSDT surveys. Bacteriological water quality has a strong impact on the patient outcome. The acceptable level of ET of dialysis fluid should be <0.1 EU/ml based on the results of JSDT survey. The excellent water quality in CDDS might be one of the important factors which help good patient survival in chronic dialysis in Japan.

Incidence of fever of unknown origin and subsequent antitubercular medications in hemodialysis patients: a two-year prospective study.

BACKGROUND: Hemodialysis (HD) patients are susceptible to atypical tuberculosis (TB), especially among patients presenting with fever of unknown origin (FUO), because of their impaired cellular immunity. Diagnostic trials of anti-TB drugs are therefore recommended in some TB endemic countries, including Japan, though clinical evidence for this therapy is scarce. METHODS: We prospectively collected data for incident cases of clinical FUO for two years in 78 of 169 dialysis facilities in Aichi prefecture, located in central Japan. Clinical FUO was defined as sustained fever without any localizing signs and no infiltration on chest x-rays after a one-week antibiotic trial. The baseline characteristics, subsequent body temperatures on the days of HD therapy, and names of antibiotics including anti-TB drugs with the durations of medication were reported until fever alleviation or fever sustainment for over eight weeks. RESULTS: We identified 15 newly developed clinical FUO patients among 8,125 HD patients. The incidence rate was estimated to be 92 (95% CI, 26-158) per 100,000 person-years. This corresponds to 244 cases per year among 264,473 HD patients in Japan. Anti-TB drugs were secondarily prescribed in 8 of 15 clinical FUO patients (53%). No improved fever alleviation was observed when anti-TB drugs were secondarily prescribed compared with cases in which other antibiotics were preferred. CONCLUSION: We investigated the incidence of FUO in HD patients and found that the rate was not very high, whereas anti-TB drugs were frequently used for FUO cases. The efficacy of this diagnostic therapy should be elucidated in large-scale studies.

The most recent trends of peritoneal dialysis in Japan.

By 31 December 2006, more than 260,000 patients were on dialysis therapy in Japan, and 9,243 of them (3.6%) were on peritoneal dialysis (PD). The mean age of PD patients was 5 years less than that of all patients with chronic kidney disease, and the prevalence of diabetes among them was 9.4% lower. Among the PD patients, 18.0% were being treated with a combination of PD and hemodialysis, and 33.4% were being treated with automated PD. A peritoneal equilibration test (PET) had been performed in 22.0% of the patients, who were then classified into the four PET categories. Of the tested patients, 10.3% were placed in the low (L) category; 38.3% in the low-average (LA) category; 38.4%, in the high-average (HA) category; and 13.0%, in the high (H) category. Icodextrin was used by 27.3% of patients classified L, 30.5% of those classified LA, 47.1% of those classified HA, and 55.1% of those classified H. The annual death rate was 6.1%, which was lower than the rate for HD (9.7%); and the annual withdrawal rate was 19.8%. Infectious complications were the main cause for lowered rates of patient and technique survival. Use of PD for older and diabetic patients and of combination therapy with hemodialysis are key perspectives in the most recent report on PD in Japan.

Thiamine status in end-stage chronic kidney disease patients: a single-center study.

BACKGROUND: Reportedly, thiamine deficiency, resulting from malnutrition and long-term diuretic therapy, is observed in patients with chronic kidney disease (CKD). The risk of thiamine deficiency might be enhanced, especially in end-stage CKD patients. Here, we assessed thiamine status in incident dialysis patients. METHODS: This study was a single-center cross-sectional study which included 288 consecutive patients initiated into dialysis between April 2013 and March 2017 at our hospital. Thiamine status was evaluated by high-performance liquid chromatography of whole blood samples. We evaluated the association between blood thiamine concentration and other clinical parameters. RESULTS: Of the 288 patients, 21 patients receiving thiamine supplementation at the time of dialysis initiation and 26 patients without blood thiamine measurements were excluded. In 30 patients (12.4%), blood thiamine concentration was lower than the lower limit of normal (21.3 ng/mL; dotted line). Blood thiamine concentration correlated with age, body mass index, and Barthel index (BI) score (p = 0.008, 0.012 and 0.009, respectively). Stepwise multivariate regression analysis indicated that BI scores were independent risk factors for thiamine deficiency (ß coefficients = 0.169, p = 0.013). CONCLUSIONS: The proportion of end-stage CKD patients with low blood thiamine concentration is high. Low physical function (low BI score) is an independent risk factor of thiamine deficiency. Clinicians should be aware of thiamine deficiency in end-stage CKD patients, especially those with low physical function.

Overview of regular dialysis treatment in Japan (as of 31 December 2005).

A statistical survey conducted at the end of 2005 covered 3985 medical facilities across Japan, and 3940 facilities (98.87%) responded. The dialysis population in Japan at the end of 2005 was 257,765, which showed an increase of 9599 patients (3.87%) from the end of the previous year. The number of patients per million was 2017.6. The crude death rate for one year (from the end of 2004 to the end of 2005) was 9.5%. The mean age of the patients who began dialysis (in 2005) was 66.2 years, and the mean age of the entire dialysis population was 63.9 years. The primary diseases of the patients who began dialysis were diabetic renal disease (42.0%) and chronic glomerulonephritis (27.3%). The mean (+/-SD) serum ferritin concentration of all the dialysis patients was 191 (+/-329) ng/mL. The percentages of antihypertensive agents administered to the hemodialysis patients were as follows: calcium-channel blocker, 50.3%; angiotensin-converting enzyme inhibitor, 11.5%; and angiotensin II-receptor blocker, 33.9%. Of the peritoneal dialysis patients, 33.4% used automated peritoneal dialysis devices. Moreover, 7.3% of the peritoneal dialysis patients received dialysis treatment only in the daytime, and 15% received the treatment only at night. Icodextrin solution was used by 37.2% of the peritoneal dialysis patients. The average amount of dialysis solution used by the peritoneal dialysis patients was 7.43 (+/-2.52) L/day and the average amount of removal fluid was 0.81 (+/-0.60) L/day. A peritoneal equilibration test was conducted on 67% of the patients, and the mean dialysate to plasma creatinine ratio was 0.65 (+/-0.13). The annual incidence of peritonitis in the peritoneal dialysis patients was 19.7%. Of the 126 040 patients who responded to the inquiry of the therapeutic situation of peritoneal dialysis, 676 (0.7%) had a history of encapsulated peritoneal sclerosis and 66 (0.1%) were treated for encapsulated peritoneal sclerosis. The mean life expectancy of the dialysis population in 2003 was calculated according to sex and age. Results showed that the mean life expectancy of the dialysis population was approximately 40-60% of that of the general population of the same sex and age.

Crystal structures and ligand binding of PurM proteins from Thermus thermophilus and Geobacillus kaustophilus.

Crystal structures of 5-aminoimidazole ribonucleotide (AIR) synthetase, also known as PurM, from Thermus thermophilus (Tt) and Geobacillus kaustophilus (Gk) were determined. For TtPurM, the maximum resolution was 2.2 Å and the space group was P21212 with four dimers in an asymmetric unit. For GkPurM, the maximum resolution was 2.2 Å and the space group was P21212 with one monomer in asymmetric unit. The biological unit is dimer for both TtPurM and GkPurM and the dimer structures were similar to previously determined structures of PurM in general. For TtPurM, ∼50 residues at the amino terminal were disordered in the crystal structure whereas, for GkPurM, the corresponding region covered the ATP-binding site forming an α helix in part, suggesting that the N-terminal region of PurM changes its conformation upon binding of ligands. FGAM binding site was predicted by the docking simulation followed by the MD simulation based on the SO4 (2-) binding site found in the crystal structure of TtPurM.