Epidemiologic Trends and Distributions of Imported Infectious Diseases Among Travelers to Japan Before and During the COVID-19 Pandemic, 2016 to 2021: A Descriptive Study.

BACKGROUND: Little is known about the trends of imported infectious diseases among travelers to non-endemic countries during the novel coronavirus disease 2019 (COVID-19) pandemic. This article aimed to describe those among travelers to Japan. METHODS: This is a descriptive study based on national surveillance data. Imported infectious disease cases were defined as those with a reported overseas source of infection among 15 diseases pre-selected based on the probability and impact of importation. The number of notified cases from April 2016 to March 2021 were described by disease and time of diagnosis. The relative ratio and absolute difference in case counts-both by number and per arrival-were calculated by disease comparing those from the pandemic period (April 2020-March 2021) to the pre-pandemic period (April 2016-March 2020). RESULTS: A total of 3,524 imported infectious disease cases were diagnosed during the study period, including 3,439 cases before and 85 cases during the pandemic. The proportionate distribution of diseases changed but notification counts of all 15 diseases decreased during the pandemic. Accounting for arrivals, however, seven diseases showed a two-fold or greater increase, with a notable absolute increase per million arrivals for amebiasis (60.1; 95% confidence interval [CI], 41.5-78.7), malaria (21.7; 95% CI, 10.5-33.0), and typhoid fever (9.3; 95% CI, 1.9-16.8). CONCLUSION: The epidemiology of imported infectious diseases changed during the pandemic. While the number of imported infectious disease cases decreased, the number of cases per arrivals increased considerably both in relative and absolute terms for several diseases of public health and clinical importance.

Nationwide epidemiologic and genetic surveillance of hepatitis E in Japan, 2014-2021.

Hepatitis E virus (HEV) is an emerging causative agent of acute hepatitis. To clarify the epidemiology of HEV and characterize the genetic diversity of the virus in Japan, nationwide enhanced surveillance and molecular characterization studies of HEV in Japan were undertaken from 2014 to 2021. In total, 2770 hepatitis E cases were reported, of which 88% were domestic cases, while only 4.1% represented cases following infection abroad. In addition, 57% of domestic infections occurred in males aged in their 40s-70s. For domestic cases, infection via pork meat consumption continued to be the most reported route. Analysis of the 324 sequences detected between 2016 and 2021 showed that the majority of domestic HEV strains belong to Genotype 3a (G3a) and G3b. In contrast, six of eight cases of G1 HEV reflected infection abroad. Our results suggest that HEV is circulating widely in Japan, with genotypes G3a and G3b being most prevalent. Continued surveillance is necessary to monitor future trends and changes in the epidemiology of HEV in Japan.

Trends in CD4+ Cell Counts, Viral Load, Treatment, Testing History, and Sociodemographic Characteristics of Newly Diagnosed HIV Patients in Osaka, Japan, From 2003 through 2017: A Descriptive Study.

BACKGROUND: The CD4 cell count of patients during diagnosis and distribution of CD4 cell counts in the patient population are important to understand infection-diagnosis interval and incidence rate of human immunodeficiency virus (HIV) infection, respectively. However, this information has not been published in Japan. This study aimed to describe the change in CD4 cell count trends and clarify the change in patients' characteristics in association with the CD4 cell count information. METHODS: A descriptive study was conducted to analyze the medical records of patients with HIV who visited one of the largest acquired immunodeficiency syndrome (AIDS) core hospitals in western Japan. The basic characteristics, CD4 cell counts, viral loads, and diagnosis-treatment intervals between the first (2003-2010) and second (2011-2017) halves of the study duration were compared. RESULTS: The distribution of CD4 cell counts significantly changed between 2003-2010 and 2011-2017 (χ2 = 20.42, P < 0.001). The proportion of CD4 cell count <200 cells/mm3 increased (38.8% in 2003 to 45.9% in 2017), whereas CD4 cell count ≥500 cells/mm3 decreased (19.4% in 2003 to 12.2% in 2017). Moreover, the distributions of age groups, history of HIV screening test, patient outcomes, HIV viral load, and diagnosis-treatment interval also significantly changed (χ2 = 25.55, P < 0.001; χ2 = 8.37, P = 0.015; χ2 = 6.07, P = 0.014; χ2 = 13.36, P = 0.020; χ2 = 173.76, P < 0.001, respectively). CONCLUSION: This study demonstrated the fundamental trends of the HIV epidemic in Osaka, Japan between 2003-2010 and 2011-2017 and indicated that the incidence rate of HIV was decreasing in Japan.

Excess All-Cause Deaths during Coronavirus Disease Pandemic, Japan, January-May 20201.

To provide insight into the mortality burden of coronavirus disease (COVID-19) in Japan, we estimated the excess all-cause deaths for each week during the pandemic, January-May 2020, by prefecture and age group. We applied quasi-Poisson regression models to vital statistics data. Excess deaths were expressed as the range of differences between the observed and expected number of all-cause deaths and the 95% upper bound of the 1-sided prediction interval. A total of 208-4,322 all-cause excess deaths at the national level indicated a 0.03%-0.72% excess in the observed number of deaths. Prefecture and age structure consistency between the reported COVID-19 deaths and our estimates was weak, suggesting the need to use cause-specific analyses to distinguish between direct and indirect consequences of COVID-19.

Clinical Characteristics and Risk Factors for Severe Outcomes of Novel Coronavirus Infection, January-March 2020, Japan.

BACKGROUND: Notifications of novel coronavirus infections increased in early 2020 in Japan. We described characteristics of novel coronavirus infection cases and analyzed risk factors for severe outcomes. METHODS: Cases were persons with laboratory-confirmed novel coronavirus infection reported under national surveillance between January and March 2020. Clinical characteristics were described, and risk factors of (1) intensive care unit [ICU] admission and (2) invasive ventilation/death were analyzed using Poisson regression. RESULTS: Among the 516 cases analyzed, median age was 60 years (range: 1-97 years) and 285 (55%) were male. Common symptoms/signs were fever (375/475, 79%), cough (353/465, 76%), and pneumonia (245/387, 63%). Ten (2%) cases died. Of the 348 cases with data, 50 (14%) required invasive ventilation. Adjusted for each other, male gender and 1-year increase in age were associated with ICU admission (risk ratio [RR] 4.18; 95% confidence interval [CI], 1.69-10.32 and RR 1.05; 95% CI, 1.03-1.08, respectively) and invasive ventilation/death (RR 2.79; 95% CI, 1.49-5.21 and RR 1.06; 95% CI, 1.04-1.08, respectively). Diabetes, dyslipidemia, hyperuricemia, and lung diseases were also associated with severe outcomes. Of the 80 cases asymptomatic at hospitalization, 40 developed symptoms and five of them >70 years of age required invasive ventilation. CONCLUSIONS: The early stage of the novel coronavirus epidemic in Japan disproportionately affected the elderly. Older age, male gender, and underlying conditions were associated with severe outcomes. Notably, some elderly case-patients who were asymptomatic at diagnosis and promptly hospitalized still went on to develop severe disease, indicating the importance of careful monitoring of certain populations.

Clinical Isolates of Human Coronavirus 229E Bypass the Endosome for Cell Entry.

Human coronavirus 229E (HCoV-229E), a causative agent of the common cold, enters host cells via two distinct pathways: one is mediated by cell surface proteases, particularly transmembrane protease serine 2 (TMPRSS2), and the other by endosomal cathepsin L. Thus, specific inhibitors of these proteases block virus infection. However, it is unclear which of these pathways is actually utilized by HCoV-229E in the human respiratory tract. Here, we examined the mechanism of cell entry used by a pseudotyped virus bearing the HCoV-229E spike (S) protein in the presence or absence of protease inhibitors. We found that, compared with a laboratory strain isolated in 1966 and passaged for a half century, clinical isolates of HCoV-229E were less likely to utilize cathepsin L; rather, they showed a preference for TMPRSS2. Two amino acid substitutions (R642M and N714K) in the S protein of HCoV-229E clinical isolates altered their sensitivity to a cathepsin L inhibitor, suggesting that these amino acids were responsible for cathepsin L use. After 20 passages in HeLa cells, the ability of the isolate to use cathepsin increased so that it was equal to that of the laboratory strain; this increase was caused by an amino acid substitution (I577S) in the S protein. The passaged virus showed a reduced ability to replicate in differentiated airway epithelial cells cultured at an air-liquid interface. These results suggest that the endosomal pathway is disadvantageous for HCoV-229E infection of human airway epithelial cells; therefore, clinical isolates are less able to use cathepsin. IMPORTANCE: Many enveloped viruses enter cells through endocytosis. Viral spike proteins drive the fusion of viral and endosomal membranes to facilitate insertion of the viral genome into the cytoplasm. Human coronavirus 229E (HCoV-229E) utilizes endosomal cathepsin L to activate the spike protein after receptor binding. Here, we found that clinical isolates of HCoV-229E preferentially utilize the cell surface protease TMPRSS2 rather than endosomal cathepsin L. The endosome is a main site of Toll-like receptor recognition, which then triggers an innate immune response; therefore, HCoV-229E presumably evolved to bypass the endosome by entering the cell via TMPRSS2. Thus, the virus uses a simple mechanism to evade the host innate immune system. Therefore, therapeutic agents for coronavirus-mediated diseases, such as severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS), should target cell surface TMPRSS2 rather than endosomal cathepsin.

Comprehensive analysis of the Co-structures of dipeptidyl peptidase IV and its inhibitor.

BACKGROUND: We comprehensively analyzed X-ray cocrystal structures of dipeptidyl peptidase IV (DPP-4) and its inhibitor to clarify whether DPP-4 alters its general or partial structure according to the inhibitor used and whether DPP-4 has a common rule for inhibitor binding. RESULTS: All the main and side chains in the inhibitor binding area were minimally altered, except for a few side chains, despite binding to inhibitors of various shapes. Some residues (Arg125, Glu205, Glu206, Tyr662 and Asn710) in the area had binding modes to fix a specific atom of inhibitor to a particular spatial position in DPP-4. We found two specific water molecules that were common to 92 DPP-4 structures. The two water molecules were close to many inhibitors, and seemed to play two roles: maintaining the orientation of the Glu205 and Glu206 side chains through a network via the water molecules, and arranging the inhibitor appropriately at the S2 subsite. CONCLUSIONS: Our study based on high-quality resources may provide a necessary minimum consensus to help in the discovery of a novel DPP-4 inhibitor that is commercially useful.

The host protease TMPRSS2 plays a major role in in vivo replication of emerging H7N9 and seasonal influenza viruses.

UNLABELLED: Proteolytic cleavage of the hemagglutinin (HA) protein is essential for influenza A virus (IAV) to acquire infectivity. This process is mediated by a host cell protease(s) in vivo. The type II transmembrane serine protease TMPRSS2 is expressed in the respiratory tract and is capable of activating a variety of respiratory viruses, including low-pathogenic (LP) IAVs possessing a single arginine residue at the cleavage site. Here we show that TMPRSS2 plays an essential role in the proteolytic activation of LP IAVs, including a recently emerged H7N9 subtype, in vivo. We generated TMPRSS2 knockout (KO) mice. The TMPRSS2 KO mice showed normal reproduction, development, and growth phenotypes. In TMPRSS2 KO mice infected with LP IAVs, cleavage of HA was severely impaired, and consequently, the majority of LP IAV progeny particles failed to gain infectivity, while the viruses were fully activated proteolytically in TMPRSS2+/+ wild-type (WT) mice. Accordingly, in contrast to WT mice, TMPRSS2 KO mice were highly tolerant of challenge infection by LP IAVs (H1N1, H3N2, and H7N9) with ≥1,000 50% lethal doses (LD50) for WT mice. On the other hand, a high-pathogenic H5N1 subtype IAV possessing a multibasic cleavage site was successfully activated in the lungs of TMPRSS2 KO mice and killed these mice, as observed for WT mice. Our results demonstrate that recently emerged H7N9 as well as seasonal IAVs mainly use the specific protease TMPRSS2 for HA cleavage in vivo and, thus, that TMPRSS2 expression is essential for IAV replication in vivo. IMPORTANCE: Influenza A virus (IAV) is a leading pathogen that infects and kills many humans every year. We clarified that the infectivity and pathogenicity of IAVs, including a recently emerged H7N9 subtype, are determined primarily by a host protease, TMPRSS2. Our data showed that TMPRSS2 is the key host protease that activates IAVs in vivo through proteolytic cleavage of their HA proteins. Hence, TMPRSS2 is a good target for the development of anti-IAV drugs. Such drugs could also be effective for many other respiratory viruses, including the recently emerged Middle East respiratory syndrome (MERS) coronavirus, because they are also activated by TMPRSS2 in vitro. Consequently, the present paper could have a large impact on the battle against respiratory virus infections and contribute greatly to human health.

Enterohemorrhagic Escherichia coli outbreaks related to childcare facilities in Japan, 2010-2013.

BACKGROUND: Enterohemorrhagic Escherichia coli (EHEC) is an important cause of gastroenteritis in Japan. Although non-O157 EHEC infections have been increasingly reported worldwide, their impact on children has not been well described. METHODS: We collected national surveillance data of EHEC infections reported between 2010 and 2013 in Japan and characterized outbreaks that occurred in childcare facilities. Per Japanese outbreak investigation protocol, faecal samples from contacts of EHEC cases were collected regardless of symptomatic status. Cases and outbreaks were described by demographics, dates of diagnosis and onset, clinical manifestations, laboratory data, and relation to specific outbreaks in childcare facilities. RESULTS: During 2010-2013, a total of 68 EHEC outbreaks comprised of 1035 cases were related to childcare facilities. Among the 66 outbreaks caused by a single serogroup, 29 were serogroup O26 and 22 were O157; 35 outbreaks were caused by stx1-producing strains. Since 2010, the number of reported outbreaks steadily increased, with a rise in cases and outbreaks caused by stx1-producing O26. Of 7069 EHEC cases reported nationally in 2010-2011, the majority were caused by O157 (n = 4938), relative to O26 (n = 1353) and O111 (n = 195). However, relative to 69 cases of O157 (2%) associated with childcare facility EHEC outbreaks, there were 131 (10%) such cases of O26, and this trend intensified in 2012-2013 (O157, 3%; O26, 24%; O111, 48%). Among family members of childcare facility cases, the proportion of cases that were symptomatic declined with age; 10/16 cases (63%) aged 6 years or younger, 16/53 cases (30%) 6-19 years old, 23/120 cases (19%) 20-49 years old and 2/28 cases (7%) 50 years or older were symptomatic. Thirty one of the 68 outbreaks (46%) were classified as foodborne-related. CONCLUSIONS: Childcare facility EHEC outbreaks due to non-O157 serogroups, particularly O26 and O111, increased during 2010-2013. These facilities should pay extra attention to health conditions in children. As older family members of childcare facility cases appear to be less symptomatic, they should be vigilant about hand-washing to prevent further transmission.

TMPRSS2 is an activating protease for respiratory parainfluenza viruses.

Here, we show that human parainfluenza viruses and Sendai virus (SeV), like other respiratory viruses, use TMPRSS2 for their activation. The membrane fusion proteins of respiratory viruses often possess serine and glutamine residues at the P2 and P3 positions, respectively, but these residues were not critical for cleavage by TMPRSS2. However, mutations of these residues affected SeV growth in specific epithelial cell lines, suggesting the importance of these residues for SeV replication in epithelia.

Biochemical analysis of packing and assembling heptad repeat motifs in the coronavirus spike protein trimer.

During a coronavirus infection, the spike protein undergoes sequential structural transitions triggered by its receptor and the host protease at the interface between the virus and cell membranes, thereby mediating membrane fusion. After receptor binding, the heptad repeat motif (HR1/HR2) within the viral spike protein bridges the viral and cellular membranes; however, the intermediate conformation adopted by the spike protein when drawing the viral and cellular membranes into close proximity remains unclear due to its transient and unstable nature. Here, we experimentally induced conformational changes in the spike protein of a murine coronavirus by incubating the virus with its receptor, followed by exposure to trypsin. We then treated the virus/receptor complex with proteinase K to probe the tightly packed core structure of the spike protein. The conformations of the spike protein were predicted from the sizes of the protease digestion products detected by western blot analysis. Upon receptor binding, two bands (each showing different reactivity with a fusion-inhibiting HR2-peptide) were detected; we propose that these bands correspond to the packed and unpacked HR1/HR2 motifs. After trypsin-mediated triggering, measurement of temperature and time dependency revealed that packing of the remaining unpacked HR1/HR2 motifs and assembly of three HR1 motifs in a trimer occur almost simultaneously. Thus, the trimeric spike protein adopts an asymmetric-unassembled conformation after receptor binding, followed by direct assembly into the post-fusion form triggered by the host protease. This biochemical study provides mechanistic insight into the previously unknown intermediate structure of the viral fusion protein.IMPORTANCEDuring infection by an enveloped virus, receptor binding triggers fusion between the cellular membrane and the virus envelope, enabling delivery of the viral genome to the cytoplasm. The viral spike protein mediates membrane fusion; however the molecular mechanism underlying this process is unclear. This is because using structural biology methods to track the transient conformational changes induced in the unstable spike trimer is challenging. Here, we harnessed the ability of protease enzymes to recognize subtle differences on protein surfaces, allowing us to detect structural differences in the spike protein before and after conformational changes. Differences in the size of the degradation products were analyzed by western blot analysis. The proposed model explaining the conformational changes presented herein is a plausible candidate that provides valuable insight into unanswered questions in the field of virology.

Respiratory syncytial virus infection notification trends and interpretation of the reported case data, 2018-2021, Japan.

In Japan, as elsewhere, the COVID-19 pandemic affected the notification trends of respiratory syncytial virus (RSV) infection. Here, we describe the epidemiological trends of RSV cases among children reported during 2018-2021 in Japan, based on the national surveillance system. Compared to 2018 and 2019, 2020 saw an unprecedented decrease in RSV notifications per sentinel site. However, 2021 experienced an unseasonably early and high peak in week 28 (peak week in 2018 and 2019: week 37) with a large resurgence in notifications, nationwide and across regions. Regarding age, compared to 2018 and 2019, the number and proportion of cases aged 2, 3, and ≥4-years increased substantially in 2021 but the number of cases aged <1 year decreased slightly. Furthermore, in 2021, the ratio of notifications per site from outpatient clinics to hospitals increased, suggesting a proportionate increase in clinically milder case diagnoses. Notably, RSV-attributed deaths from vital statistics also dropped substantially in 2020 and rebounded in 2021, but were fewer than in 2018 or 2019. While RSV incidence likely declined in 2020 (possibly from COVID-19 countermeasures) and increased in 2021, notifications in 2021 appeared to be associated with milder presentations. Given unpredictable RSV epidemiology, continuous monitoring and pluralistic assessments are imperative.

United3D: a protein model quality assessment program that uses two consensus based methods.

In protein structure prediction, such as template-based modeling and free modeling (ab initio modeling), the step that assesses the quality of protein models is very important. We have developed a model quality assessment (QA) program United3D that uses an optimized clustering method and a simple Cα atom contact-based potential. United3D automatically estimates the quality scores (Qscore) of predicted protein models that are highly correlated with the actual quality (GDT_TS). The performance of United3D was tested in the ninth Critical Assessment of protein Structure Prediction (CASP9) experiment. In CASP9, United3D showed the lowest average loss of GDT_TS (5.3) among the QA methods participated in CASP9. This result indicates that the performance of United3D to identify the high quality models from the models predicted by CASP9 servers on 116 targets was best among the QA methods that were tested in CASP9. United3D also produced high average Pearson correlation coefficients (0.93) and acceptable Kendall rank correlation coefficients (0.68) between the Qscore and GDT_TS. This performance was competitive with the other top ranked QA methods that were tested in CASP9. These results indicate that United3D is a useful tool for selecting high quality models from many candidate model structures provided by various modeling methods. United3D will improve the accuracy of protein structure prediction.

Reduced mortality during the COVID-19 outbreak in Japan, 2020: a two-stage interrupted time-series design.

BACKGROUND: Coronavirus disease 2019 (COVID-19) continues to be a major global health burden. This study aims to estimate the all-cause excess mortality occurring in the COVID-19 outbreak in Japan, 2020, by sex and age group. METHODS: Daily time series of mortality for the period January 2015-December 2020 in all 47 prefectures of Japan were obtained from the Ministry of Health, Labour and Welfare, Japan. A two-stage interrupted time-series design was used to calculate excess mortality. In the first stage, we estimated excess mortality by prefecture using quasi-Poisson regression models in combination with distributed lag non-linear models, adjusting for seasonal and long-term variations, weather conditions and influenza activity. In the second stage, we used a random-effects multivariate meta-analysis to synthesize prefecture-specific estimates at the nationwide level. RESULTS: In 2020, we estimated an all-cause excess mortality of -20 982 deaths [95% empirical confidence intervals (eCI): -38 367 to -5472] in Japan, which corresponded to a percentage excess of -1.7% (95% eCI: -3.1 to -0.5) relative to the expected value. Reduced deaths were observed for both sexes and in all age groups except those aged <60 and 70-79 years. CONCLUSIONS: All-cause mortality during the COVID-19 outbreak in Japan in 2020 was decreased compared with a historical baseline. Further evaluation of cause-specific excess mortality is warranted.

Epidemiology of Coronavirus Disease 2019 in Japan: Descriptive Findings and Lessons Learned through Surveillance during the First Three Waves.

INTRODUCTION: Coronavirus disease 2019 (COVID-19) has caused unprecedented global morbidity and mortality. Japan has faced three epidemic "waves" of COVID-19 from early 2020 through early 2021. Here we narratively review the three waves in Japan, describe the key epidemiologic features of COVID-19, and discuss lessons learned. METHODS: We assessed publicly available surveillance data, routine surveillance reports, and other relevant sources-multiple indicators were monitored to improve interpretation of surveillance data. Weekly trends for each wave were described based on the number of case notifications; number of tests performed; proportion of those tests that were positive for the novel coronavirus; the prevalent number of COVID-19 hospitalizations (total hospitalizations and those categorized as severe); and number of COVID-19 deaths. For each indicator and wave, we recorded the first calendar week to show an increase over two consecutive previous weeks, along with the peak week. RESULTS: The spring wave was characterized by detection of cases imported from China, followed by notifications of sporadic cases without travel history, clusters, and mild/asymptomatic cases. The summer wave saw a large increase in notifications and a younger age distribution, but in the context of increased testing with lower test positivity. The winter wave brought considerable morbidity and mortality, surpassing the cumulative case counts and fatalities from the earlier waves, with high peak values. Overall, relative to the first wave, the burden of severe outcomes was lower in the second and higher in the third wave, but varied by prefecture. In all three waves, severe outcomes peaked after notification counts and test positivity peaked; severe outcomes were also consistently skewed toward the elderly. CONCLUSIONS: Important lessons were learned from each wave and across waves-some aspects remained constant, while others changed over time. In order to rapidly detect an increase in incidence, continuous, timely, and sensitive surveillance-using multiple information sources with careful interpretations-will be key in COVID-19 control.

Method for predicting homology modeling accuracy from amino acid sequence alignment: the power function.

We have devised a power function (PF) that can predict the accuracy of a three-dimensional (3D) structure model of a protein using only amino acid sequence alignments. This Power Function (PF) consists of three parts; (1) the length of a model, (2) a homology identity percent value and (3) the agreement rate between PSI-PRED secondary structure prediction and the secondary structure judgment of a reference protein. The PF value is mathematically computed from the execution process of homology search tools, such as FASTA or various BLAST programs, to obtain the amino acid sequence alignments. There is a high correlation between the global distance test-total score (GDT_TS) value of the protein model based upon the PF score and the GDT_TS(MAX) value used as an index of protein modeling accuracy in the international contest Critical Assessment of Techniques for Protein Structure Prediction (CASP). Accordingly, the PF method is valuable for constructing a highly accurate model without wasteful calculations of homology modeling that is normally performed by an iterative method to move the main chain and side chains in the modeling process. Moreover, a model with higher accuracy can be obtained by combining the models ordered by the PF score with models sorted by the size of the CIRCLE score. The CIRCLE software is a 3D-1D program, in which energetic stabilization is estimated based upon the experimental environment of each amino acid residue in the protein solution or protein crystals.

HUMAN FAMSD-BASE: high quality protein structure model database for the human genome using the FAMSD homology modeling method.

Almost all proteins express their biological functions through the structural conformation of their specific amino acid sequences. Therefore, acquiring the three-dimensional structures of proteins is very important to elucidate the role of a particular protein. We had built protein structure model databases, which is called RIKEN FAMSBASE (http://famshelp.gsc.riken.jp/famsbase/). The RIKEN FAMSBASE is a genome-wide protein structure model database that contains a large number of protein models from many organisms. The HUMAN FAMSBASE that is one part of the RIKEN FAMSBASE contains many protein models for human genes, which are significant in the pharmaceutical and medicinal fields. We have now implemented an update of the human protein modeling database consisting of 242918 constructed models against the number of 20743 human protein sequences with an improved modeling method called Full Automatic protein Modeling System Developed (FAMSD). The results of our benchmark test of the FAMSD method indicated that it has an excellent capability to pack amino acid side-chains with correct torsion angles in addition to the main-chain, while avoiding the formation of atom-atom collisions that are not found in experimental structures. This new protein structure model database for human genes, which is named HUMAN FAMSD-BASE, is open to the public as a component part of the RIKEN FAMSBASE at http://mammalia.gsc.riken.jp/human_famsd/. A significant improvement of the HUMAN FAMSD-BASE in comparison with the preceding HUMAN FAMSBASE was verified in the benchmark test of this paper. The HUMAN FAMSD-BASE will have an important impact on the progress of biological science.

New protein structure model evaluation methods that include a side-chain consensus score for the protein modeling.

Selecting the best quality model from a set of predicted structures is one of the most important aspects of protein structure prediction. We have developed model quality assessment programs that select high quality models which account for both the Calpha backbone and side-chain atom positions. The new methods are based on the consensus method with consideration of the side-chain environment of a protein structure and the secondary structure agreement. This Side-chain Environment Consensus (SEC) method is compared with the conventional consensus method, 3D-Jury (Ginalski K. et al., Bioinformatics, 19, 1015-1018 (2003)), which takes into account only the Calpha backbone atoms of the protein model. As the result, it was found that the SEC method selects the models with more accurate positioning of the side-chain atoms than the 3D-Jury method. When the SEC method was used in combination with the 3D-Jury method (3DJ+SEC), models were selected with improved quality both in the Calpha backbone and side-chain atom positions. Moreover, the CIRCLE (CCL) method (Terashi G. et al., Proteins, 69 (Suppl. 8), 98-107 (2007)) based on the 3D-1D profile score has been shown to select the best possible models that are the closest to the native structures from candidate models. Accordingly, the 3DJ+SEC+CCL method, in which CIRCLE is used after reducing the number of candidates by the 3DJ+SEC consensus method, was found to be very effective in selecting high quality models. Thus, the best method (the 3DJ+SEC+CCL method) includes the consensus approaches of the Calpha backbone and the side-chains, the secondary structure agreement and the 3D-1D profile score which corresponds to the free energy-like score in the residues of the protein model. In short, new algorithms are introduced in protein structure evaluation methods that are based on a side-chain consensus score. Additionally, in order to apply the 3DJ+SEC+CCL method and indicate the usefulness of this method, a model of human Cabin1, a protein associated with p53 function and cancer, is created using various internet modeling and alignment servers.

Dynamic influence of the two membrane-proximal immunoglobulin-like domains upon the peptide-binding platform domain in class I and class II major histocompatibility complexes: normal mode analysis.

Major histocompatibility complexes (MHCs) mainly fall into class I and class II. The two classes have similar structures, with two membrane-proximal immunoglobulin-like domains and a peptide-binding platform domain, though their organizations are different. We simulated the dynamics of a whole and partial model deficient in either of the two membrane-proximal domains for class I and class II using normal mode analysis. Our study showed that the influence of the two membrane-proximal domains upon the dynamics of the platform domain were decisively different between class II and class I. Both membrane-proximal domains (the alpha2 and beta2 domains) of class II MHC, especially the alpha2 domain, influenced the most important pocket that accommodates a large hydrophobic anchor side chain of the N-terminal side of the bound peptide, though the pocket was not in the alpha2 domain neighborhood. By contrast, the two membrane-proximal domains (the alpha3 and beta2m domains) of class I MHC had little influence upon the most important pocket that accommodates the N-terminal residue of the bound peptide. These results suggest that the two membrane-proximal domains of class II MHC have a greater influence upon peptide-binding than those of class I MHC.

FAMSD: A powerful protein modeling platform that combines alignment methods, homology modeling, 3D structure quality estimation and molecular dynamics.

The prediction of a protein three-dimensional (3D) structure is one of the most important challenges in computational structural biology. We have developed an automatic protein 3D structure prediction method called FAMSD. FAMSD is based on a comparative modeling method which consists of the following four steps: (1) generating and selecting sequence alignments between target and template proteins; (2) constructing 3D structure models based on each selected alignment; (3) selecting the best 3D structure model and (4) refining the selected model. In the FAMSD method, sequence alignment programs such as a series of BLAST programs, SP3 and SPARKS2 programs, the homology modeling program FAMS (Full Automatic Modeling System), the model quality estimation program CIRCLE and the molecular dynamics program APRICOT were used in combination to construct high quality protein models. To assess the FAMSD method we have participated in the 8th Critical Assessment of Techniques for Protein Structure Prediction (CASP8) experiment. The results of our original assessment indicate that the FAMSD method offers excellent capability in packing side-chains with the correct torsion angles while avoiding the formation of atom-atom collisions. Since side-chain packing plays a significant role in defining the biological function of proteins, this method is a valuable resource in biological, pharmaceutical and medicinal research efforts.