Production of lactulose from lactose using a novel cellobiose 2-epimerase from Clostridium disporicum.

Lactulose is a semisynthetic nondigestive sugar derived from lactose, with wide applications in the food and pharmaceutical industries. Its biological production routes which use cellobiose 2-epimerase (C2E) as the key enzyme have attracted widespread attention. In this study, a set of C2Es from different sources were overexpressed in Escherichia coli to produce lactulose. We obtained a novel and highly efficient C2E from Clostridium disporicum (CDC2E) to synthesize lactulose from lactose. The effects of different heat treatment conditions, reaction pH, reaction temperature, and substrate concentrations were investigated. Under the optimum biotransformation conditions, the final concentration of lactulose was up to 1.45 M (496.3 g/L), with a lactose conversion rate of 72.5 %. This study provides a novel C2E for the biosynthesis of lactulose from low-cost lactose.

Generative Variational-Contrastive Learning for Self-Supervised Point Cloud Representation.

Self-supervised representation learning for 3D point clouds has attracted increasing attention. However, existing methods in the field of 3D computer vision generally use fixed embeddings to represent the latent features, and impose hard constraints on the embeddings to make the latent feature values of the positive samples converge to consistency, which limits the ability of feature extractors to generalize over different data domains. To address this issue, we propose a Generative Variational-Contrastive Learning (GVC) model, where Gaussian distribution is used to construct a continuous, smoothed representation of the latent features. A distribution constraint and cross-supervision are constructed to improve the transfer ability of the feature extractor over synthetic and real-world data. Specifically, we design a variational contrastive module to constrain the feature distribution instead of feature values corresponding to each sample in the latent space. Moreover, a generative cross-supervision module is introduced to preserve the invariance features and promote the consistency of feature distribution among positive samples. Experimental results demonstrate that GVC achieves SOTA on different downstream tasks. In particular, with only pre-training on the synthetic dataset, GVC achieves a lead of 8.4% and 14.2% when transferring to the real-world dataset in the linear classification and few-shot classification.

Metabolic engineering of Escherichia coli for efficient production of L-alanyl-L-glutamine.

BACKGROUND: L-Alanyl-L-glutamine (AQ) is a functional dipeptide with high water solubility, good thermal stability and high bioavailability. It is widely used in clinical treatment, post-operative rehabilitation, sports health care and other fields. AQ is mainly produced via chemical synthesis which is complicated, time-consuming, labor-intensive, and have a low yield accompanied with the generation of by-products. It is therefore highly desirable to develop an efficient biotechnological process for the industrial production of AQ. RESULTS: A metabolically engineered E. coli strain for AQ production was developed by over-expressing L-amino acid α-ligase (BacD) from Bacillus subtilis, and inactivating the peptidases PepA, PepB, PepD, and PepN, as well as the dipeptide transport system Dpp. In order to use the more readily available substrate glutamic acid, a module for glutamine synthesis from glutamic acid was constructed by introducing glutamine synthetase (GlnA). Additionally, we knocked out glsA-glsB to block the first step in glutamine metabolism, and glnE-glnB involved in the ATP-dependent addition of AMP/UMP to a subunit of glutamine synthetase, which resulted in increased glutamine supply. Then the glutamine synthesis module was combined with the AQ synthesis module to develop the engineered strain that uses glutamic acid and alanine for AQ production. The expression of BacD and GlnA was further balanced to improve AQ production. Using the final engineered strain p15/AQ10 as a whole-cell biocatalyst, 71.7 mM AQ was produced with a productivity of 3.98 mM/h and conversion rate of 71.7%. CONCLUSION: A metabolically engineered strain for AQ production was successfully developed via inactivation of peptidases, screening of BacD, introduction of glutamine synthesis module, and balancing the glutamine and AQ synthesis modules to improve the yield of AQ. This work provides a microbial cell factory for efficient production of AQ with industrial potential.

CMAUP: a database of collective molecular activities of useful plants.

The beneficial effects of functionally useful plants (e.g. medicinal and food plants) arise from the multi-target activities of multiple ingredients of these plants. The knowledge of the collective molecular activities of these plants facilitates mechanistic studies and expanded applications. A number of databases provide information about the effects and targets of various plants and ingredients. More comprehensive information is needed for broader classes of plants and for the landscapes of individual plant's multiple targets, collective activities and regulated biological pathways, processes and diseases. We therefore developed a new database, Collective Molecular Activities of Useful Plants (CMAUP), to provide the collective landscapes of multiple targets (ChEMBL target classes) and activity levels (in 2D target-ingredient heatmap), and regulated gene ontologies (GO categories), biological pathways (KEGG categories) and diseases (ICD blocks) for 5645 plants (2567 medicinal, 170 food, 1567 edible, 3 agricultural and 119 garden plants) collected from or traditionally used in 153 countries and regions. These landscapes were derived from 47 645 plant ingredients active against 646 targets in 234 KEGG pathways associated with 2473 gene ontologies and 656 diseases. CMAUP (http://bidd2.nus.edu.sg/CMAUP/) is freely accessible and searchable by keywords, plant usage classes, species families, targets, KEGG pathways, gene ontologies, diseases (ICD code) and geographical locations.

NPASS: natural product activity and species source database for natural product research, discovery and tool development.

There has been renewed interests in the exploration of natural products (NPs) for drug discovery, and continuous investigations of the therapeutic claims and mechanisms of traditional and herbal medicines. In-silico methods have been employed for facilitating these studies. These studies and the optimization of in-silico algorithms for NP applications can be facilitated by the quantitative activity and species source data of the NPs. A number of databases collectively provide the structural and other information of ∼470 000 NPs, including qualitative activity information for many NPs, but only ∼4000 NPs are with the experimental activity values. There is a need for the activity and species source data of more NPs. We therefore developed a new database, NPASS (Natural Product Activity and Species Source) to complement other databases by providing the experimental activity values and species sources of 35 032 NPs from 25 041 species targeting 5863 targets (2946 proteins, 1352 microbial species and 1227 cell-lines). NPASS contains 446 552 quantitative activity records (e.g. IC50, Ki, EC50, GI50 or MIC mainly in units of nM) of 222 092 NP-target pairs and 288 002 NP-species pairs. NPASS, http://bidd2.nus.edu.sg/NPASS/, is freely accessible with its contents searchable by keywords, physicochemical property range, structural similarity, species and target search facilities.

Database and Bioinformatics Studies of Probiotics.

Probiotics have been widely explored for health benefits, animal cares, and agricultural applications. Recent advances in microbiome, microbiota, and microbial dark matter research have fueled greater interests in and paved ways for the study of the mechanisms of probiotics and the discovery of new probiotics from uncharacterized microbial sources. A probiotics database named PROBIO was developed to facilitate these efforts and the need for the information on the known probiotics, which provides the comprehensive information about the probiotic functions of 448 marketed, 167 clinical trial/field trial, and 382 research probiotics for use or being studied for use in humans, animals, and plants. The potential applications of the probiotics data are illustrated by several literature-reported investigations, which have used the relevant information for probing the function and mechanism of the probiotics and for discovering new probiotics. PROBIO can be accessed free of charge at http://bidd2.nus.edu.sg/probio/homepage.htm .

Optimization of beta-carotene production by a newly isolated Serratia marcescens strain

beta-carotene is a commonly used food colorant. In this work, a novel beta-carotene producing strain, Serratia marcescens RB3, was isolated and identified by physiological and biochemical tests, as well as 16S rDNA sequence analysis. The production of beta-carotene by S. marcescens RB3 was identified through HPLC analysis. The cultivation conditions for beta-carotene production by S. marcescens RB3 were optimized as 2.0 percent lactose, 2.0 percent peptone, 0.3 percent beef extract, 1.0 percent NaCl supplemented with 0.05 percent Fe2+, pH 6.0 and 30ºC. Under the optimal conditions, the yield of beta-carotene achieved 2.45 ug/mL.

Use of the pyrG gene as a food-grade selection marker in Monascus.

Ma-pyrG was cloned from Monascus aurantiacus AS3.4384 using degenerate PCR with primers designed with an algorithm called CODEHOP, and its complete sequence was obtained by a PCR-based strategy for screening a Monascus fosmid library. Ma-pyrG encodes orotidine-5'-phosphate decarboxylase (OMPdecase), a 283-aminoacid protein with 81% sequence identity to that from Aspergillus flavus NRRL 3357. A pyrG mutant strain from M. aurantiacus AS3.4384, named UM28, was isolated by resistance to 5-fluoroorotic acid after UV mutagenesis. Sequence analysis of this mutated gene revealed that it contained a point mutation at nucleotide position +220. Plasmid pGFP-pyrG, bearing the green fluorescent protein gene (GFP) as a model gene and Ma-pyrG as a selection marker, were constructed. pGFP-pyrG were successfully transformed into UM28 by using the PEG method.

Individualized prostate biopsy strategy for Chinese patients with different prostate-specific antigen levels.

AIM: To evaluate the best individualized prostate biopsy strategies for Chinese patients with suspected prostate cancer. METHODS: The present study included 221 Chinese patients who underwent transrectal ultrasound guided prostate biopsies for the first time. All patients underwent the same 10-core biopsy protocol. In addition to the Hodge sextant technique, four more biopsies were obtained from the base and middle regions of bilateral peripheral zones. The differences between 10-core and sextant strategies in cancer detection among patients with different prostate specific antigen (PSA) levels were evaluated. The relationship between PSA level, number of positive biopsy cores and organ-confined cancer rate in prostate cancer patients was also analyzed. RESULTS: The overall prostate cancer detection rate was 40.7% in the 221 patients. The 10-core strategy increased cancer detection by 6.67% (6/90) in our patients (P < 0.05). The increased cancer detection rates decreased significantly when the patient PSA level increased from 0-20 ng/mL to 20.1-50 ng/mL and > 50 ng/mL (P < 0.01). The number of positive biopsy cores in prostate cancer patients increased significantly with increasing patient PSA level (P < 0.01). The rate of organ-confined prostate cancer decreased significantly with increasing patient PSA level (P < 0.01). CONCLUSION: The extended 10-core strategy is recommended for Chinese patients with PSA = or < 20 ng/mL and the sextant strategy is recommended for those with PSA > 50 ng/mL. For patients with PSA ranging from 20.1 ng/mL to 50 ng/mL, the 10-core strategy should be applied in patients with life expectancy = or > 10 years and the sextant strategy should be applied in those with life expectancy < 10 years.

[Evaluation of the results of fine-needle aspiration liver biopsies and the complications in 2528 cases].

OBJECTIVE: To study the value of fine-needle aspiration biopsy (FNAB) in diagnosing primary liver cancer (PLC) and its major complications. METHODS: From June 1, 1985 to May 31, 2005, 2528 patients who were presented with suspected PLC underwent ultrasound-guided FNAB in the Cancer Hospital of Fudan University. The results were retrospectively reviewed. RESULTS: Of those 2528 cases, there was malignancy in 2061 patients (81.53%), of which 1704 were diagnosed as primary liver neoplasms; 41 were diagnosed as metastatic carcinoma, and 316 were not further classified as primary or metastatic. No malignancy was found in 431 cases (17.05%). In 36 cases (1.42%), suspicious malignancy or anaplasia was suggested. Follow-up results showed that all the 2061 positive cases were verified to be malignant and there were no false positive cases. 163 of the 431 negative cases were verified to be malignant in the follow-ups, of which 136 cases were PLC; 28 of the 36 suspicious malignancy or anaplasia were proven to be malignant (all were PLC). The sensitivity, specificity, positive predictive value, negative predictive value and overall accuracy of our FNAB for diagnosing liver malignancy were 91.52%, 100.00%, 100.00%, 59.10% and 92.44%, respectively, and 81.01% cases were diagnosed by FNAB in all the 2096 cases with PLC. Cytological examinations of the smears obtained by FNAB correctly distinguished primary and secondary malignancy in 77.49% of the patients. After FNAB, 11 patients (0.44%) had intraperitoneal hemorrhages and 5 cases (0.20%) had needle tract implantation metastases. CONCLUSIONS: FNAB is important and effective for determining the malignancy potential of liver tumors, especially for PLC. Complications related to FNA were rather rare, therefore, this technique may be easily applied to clinical practice.