Journal of Microbiology

Journal Articles

Cytophaga hutchinsonii chu_2177, encoding the O-antigen ligase, is essential for cellulose degradation: Yahong Tan , Wenxia Song , Lijuan Gao , Weican Zhang , Xuemei Lu; J. Microbiol. 2022;60(4):364-374. Published online January 7, 2022; DOI: https://doi.org/10.1007/s12275-022-1531-3

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Cytophaga hutchinsonii can efficiently degrade crystalline cellulose, in which the cell surface cellulases secreted by the type IX secretion system (T9SS) play important roles, but the degradation mechanism remains unclear, and the anchor mechanism of cellulases on the outer membrane in C. hutchinsonii has not been studied. Here, chu_2177 was identified by transposon mutagenesis and was proved to be indispensable for cellulose utilization in C. hutchinsonii. Disruption of chu_2177 resulted in O-antigen deficiency and chu_ 177 could confer O-antigen ligase activity upon an Escherichia coli waal mutant, indicating that chu_2177 encoded the Ontigen ligase. Moreover, deletion of chu_2177 caused defects in cellulose utilization, cell motility, biofilm formation, and stress resistance. Further study showed that the endoglucanase activity was markedly decreased in the outer membrane but was increased in the culture fluid without chu_2177. Western blot proved that endoglucanase CHU_1336 was not located on the outer membrane but was released in the culture fluid of the Δ2177 mutant. Further proteomics analysis showed that many cargo proteins of T9SS were missing in the outer membrane of the Δ2177 mutant. Our study revealed that the deletion of chu_2177 affected the localization of many T9SS cargo proteins including cellulases on the outer membrane of C. hutchinsonii.

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Screening and genome-wide analysis of lignocellulose-degrading bacteria from humic soil
Tianjiao Zhang, Shuli Wei, Yajie Liu, Chao Cheng, Jie Ma, Linfang Yue, Yanrong Gao, Yuchen Cheng, Yongfeng Ren, Shaofeng Su, Xiaoqing Zhao, Zhanyuan Lu
Frontiers in Microbiology.2023;[Epub] CrossRef
The type IX secretion system: Insights into its function and connection to glycosylation in Cytophaga hutchinsonii
Wenxia Song, Xueke Zhuang, Yahong Tan, Qingsheng Qi, Xuemei Lu
Engineering Microbiology.2022; 2(3): 100038. CrossRef

The effects of deletion of cellobiohydrolase genes on carbon source-dependent growth and enzymatic lignocellulose hydrolysis in Trichoderma reesei: Meibin Ren , Yifan Wang , Guoxin Liu , Bin Zuo , Yuancheng Zhang , Yunhe Wang , Weifeng Liu , Xiangmei Liu , Yaohua Zhong; J. Microbiol. 2020;58(8):687-695. Published online June 10, 2020; DOI: https://doi.org/10.1007/s12275-020-9630-5

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Abstract

The saprophytic fungus Trichoderma reesei has long been used as a model to study microbial degradation of lignocellulosic biomass. The major cellulolytic enzymes of T. reesei are the cellobiohydrolases CBH1 and CBH2, which constitute more than 70% of total proteins secreted by the fungus. However, their physiological functions and effects on enzymatic hydrolysis of cellulose substrates are not sufficiently elucidated. Here, the cellobiohydrolase-encoding genes cbh1 and cbh2 were deleted, individually or combinatively, by using an auxotrophic marker-recycling technique in T. reesei. When cultured on media with different soluble carbon sources, all three deletion strains (Δcbh1, Δcbh2, and Δcbh1Δcbh2) exhibited no dramatic variation in morphological phenotypes, but their growth rates increased apparently when cultured on soluble cellulase-inducing carbon sources. In addition, Δcbh1 showed dramatically reduced growth and Δcbh1Δcbh2 could hardly grew on microcrystalline cellulose (MCC), whereas all strains grew equally on sodium carboxymethyl cellulose (CMC-Na), suggesting that the influence of the CBHs on growth was carbon source-dependent. Moreover, five representative cellulose substrates were used to analyse the influence of the absence of CBHs on saccharification efficiency. CBH1 deficiency significantly affected the enzymatic hydrolysis rates of various cellulose substrates, where acid pre-treated corn stover (PCS) was influenced the least. CBH2 deficiency reduced the hydrolysis of MCC, PCS, and acid pre-treated and delignified corncob but improved the hydrolysis ability of filter paper. These results demonstrate the specific contributions of CBHs to the hydrolysis of different types of biomass, which could facilitate the development of tailor-made strains with highly efficient hydrolysis enzymes for certain biomass types in the biofuel industry.

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An efficient CRISPR/Cas9 genome editing system based on a multiple sgRNA processing platform in Trichoderma reesei for strain improvement and enzyme production
Jiaxin Zhang, Kehang Li, Yu Sun, Cheng Yao, Weifeng Liu, Hong Liu, Yaohua Zhong
Biotechnology for Biofuels and Bioproducts.2024;[Epub] CrossRef
Transcriptome-wide analysis of a superior xylan degrading isolate Penicillium oxalicum 5–18 revealed active lignocellulosic degrading genes
Shuang Hu, Pei Han, Bao-Teng Wang, Long Jin, Hong-Hua Ruan, Feng-Jie Jin
Archives of Microbiology.2024;[Epub] CrossRef
Engineering the secretome of Aspergillus niger for cellooligosaccharides production from plant biomass
Fernanda Lopes de Figueiredo, Fabiano Jares Contesini, César Rafael Fanchini Terrasan, Jaqueline Aline Gerhardt, Ana Beatriz Corrêa, Everton Paschoal Antoniel, Natália Sayuri Wassano, Lucas Levassor, Sarita Cândida Rabelo, Telma Teixeira Franco, Uffe Hasb
Microbial Cell Factories.2024;[Epub] CrossRef
Constitutive overexpression of cellobiohydrolase 2 in Trichoderma reesei reveals its ability to initiate cellulose degradation
Yubo Wang, Meibin Ren, Yifan Wang, Lu Wang, Hong Liu, Mei Shi, Yaohua Zhong
Engineering Microbiology.2023; 3(1): 100059. CrossRef
Inducer-free recombinant protein production in Trichoderma reesei: secretory production of endogenous enzymes and heterologous nanobodies using glucose as the sole carbon source
Toshiharu Arai, Mayumi Wada, Hiroki Nishiguchi, Yasushi Takimura, Jun Ishii
Microbial Cell Factories.2023;[Epub] CrossRef
The Influence of Trctf1 Gene Knockout by CRISPR–Cas9 on Cellulase Synthesis by Trichoderma reesei with Various Soluble Inducers
Yudian Chen, Yushan Gao, Zancheng Wang, Nian Peng, Xiaoqin Ran, Tingting Chen, Lulu Liu, Yonghao Li
Fermentation.2023; 9(8): 746. CrossRef
The effect of cellobiohydrolase 1 gene knockout for composition and hydrolytic activity of the enzyme complex secreted by filamentous fungus Penicillium verruculosum
Valeriy Yu. Kislitsin, Andrey M. Chulkin, Ivan N. Zorov, Yuri А. Denisenko, Arkadiy P. Sinitsyn, Alexandra M. Rozhkova
Bioresource Technology Reports.2022; 18: 101023. CrossRef
Deciphering the efficient cellulose degradation by the thermophilic fungus Myceliophthora thermophila focused on the synergistic action of glycoside hydrolases and lytic polysaccharide monooxygenases
Xing Qin, Jiahuan Zou, Kun Yang, Jinyang Li, Xiaolu Wang, Tao Tu, Yuan Wang, Bin Yao, Huoqing Huang, Huiying Luo
Bioresource Technology.2022; 364: 128027. CrossRef

[PROTOCOL] Structural analysis of N-/O-glycans assembled on proteins in yeasts: Eun Jung Thak , Jungho Kim , Dong-Jik Lee , Jeong Yoon Kim , Hyun Ah Kang; J. Microbiol. 2018;56(1):11-23. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7468-x

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Abstract

Protein glycosylation, the most universal and diverse posttranslational modification, can affect protein secretion, stability, and immunogenicity. The structures of glycans attached to proteins are quite diverse among different organisms and even within yeast species. In yeast, protein glycosylation plays key roles in the quality control of secretory proteins, and particularly in maintaining cell wall integrity. Moreover, in pathogenic yeasts, glycans assembled on cell-surface glycoproteins can mediate their interactions with host cells. Thus, a comprehensive understanding of protein glycosylation in various yeast species and defining glycan structure characteristics can provide useful information for their biotechnological and clinical implications. Yeast-specific glycans are a target for glyco-engineering; implementing human-type glycosylation pathways in yeast can aid the production of recombinant glycoproteins with therapeutic potential. The virulenceassociated glycans of pathogenic yeasts could be exploited as novel targets for antifungal agents. Nowadays, several glycomics techniques facilitate the generation of species- and strain-specific glycome profiles and the delineation of modified glycan structures in mutant and engineered yeast cells. Here, we present the protocols employed in our laboratory to investigate the N- and O-glycan chains released from purified glycoproteins or cell wall mannoproteins in several yeast species.

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Protein Expression Platforms and the Challenges of Viral Antigen Production
Jamie R. V. Sookhoo, Zachary Schiffman, Aruna Ambagala, Darwyn Kobasa, Keith Pardee, Shawn Babiuk
Vaccines.2024; 12(12): 1344. CrossRef
Novel Botrytis cinerea Zn(II)2Cys6 Transcription Factor BcFtg1 Enhances the Virulence of the Gray Mold Fungus by Promoting Organic Acid Secretion and Carbon Source Utilization
Song Yang, Jiao Sun, Aoran Xue, Guihua Li, Chenhao Sun, Jie Hou, Qing-Ming Qin, Mingzhe Zhang
Journal of Agricultural and Food Chemistry.2024; 72(34): 18824. CrossRef
Komagataella phaffii as a Platform for Heterologous Expression of Enzymes Used for Industry
Tamara M. Khlebodarova, Natalia V. Bogacheva, Andrey V. Zadorozhny, Alla V. Bryanskaya, Asya R. Vasilieva, Danil O. Chesnokov, Elena I. Pavlova, Sergey E. Peltek
Microorganisms.2024; 12(2): 346. CrossRef
Proteomic Analysis of Cell Wall Proteins with Various Linkages in Fusarium graminearum
Heeji Moon, Kyunghun Min, Jessica Winarto, Soobin Shin, Hosung Jeon, Dae-Geun Song, Hokyoung Son
Journal of Agricultural and Food Chemistry.2024; 72(11): 6028. CrossRef
Analysis of carbohydrates and glycoconjugates by matrix‐assisted laser desorption/ionization mass spectrometry: An update for 2017–2018
David J. Harvey
Mass Spectrometry Reviews.2023; 42(1): 227. CrossRef
Rna-Seq Based Transcriptomic Analysis of the Non-Conventional Yeast Spathaspora Passalidarum During Melle-Boinot Cell Recycle in Xylose-Glucose Mixtures
Thiago Neitzel, Cleilton Santos Lima, Eduardo Hafemann, Douglas Antonio Alvaredo Paixão, Joaquim Martins Junior, Gabriela Felix Persinoti, Leandro Vieira dos Santos, jaciane ienczak
SSRN Electronic Journal .2022;[Epub] CrossRef
RNA-seq based transcriptomic analysis of the non-conventional yeast Spathaspora passalidarum during Melle-boinot cell recycle in xylose-glucose mixtures
Thiago Neitzel, Cleilton Santos Lima, Eduardo Hafemann, Douglas Antonio Alvaredo Paixão, Joaquim Martins Junior, Gabriela Felix Persinoti, Leandro Vieira dos Santos, Jaciane Lutz Ienczak
Renewable Energy.2022; 201: 486. CrossRef
Pushing and pulling proteins into the yeast secretory pathway enhances recombinant protein secretion
Richard J. Zahrl, Roland Prielhofer, Özge Ata, Kristin Baumann, Diethard Mattanovich, Brigitte Gasser
Metabolic Engineering.2022; 74: 36. CrossRef
Extension of O -Linked Mannosylation in the Golgi Apparatus Is Critical for Cell Wall Integrity Signaling and Interaction with Host Cells in Cryptococcus neoformans Pathogenesis
Eun Jung Thak, Ye Ji Son, Dong-Jik Lee, Hyunah Kim, Jung Ho Kim, Su-Bin Lee, Yu-Byeong Jang, Yong-Sun Bahn, Connie B. Nichols, J. Andrew Alspaugh, Hyun Ah Kang, Michael Lorenz
mBio.2022;[Epub] CrossRef
Metabolic labeling of glycans with isotopic glucose for quantitative glycomics in yeast
Ji-Yeon Kim, Woo Hong Joo, Dong-Soo Shin, Yong-Ill Lee, Chin Fen Teo, Jae-Min Lim
Analytical Biochemistry.2021; 621: 114152. CrossRef
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Cell Discovery.2021;[Epub] CrossRef
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Özge Ata, Burcu Gündüz Ergün, Patrick Fickers, Lina Heistinger, Diethard Mattanovich, Corinna Rebnegger, Brigitte Gasser
FEMS Yeast Research.2021;[Epub] CrossRef
Water Kefir and Derived Pasteurized Beverages Modulate Gut Microbiota, Intestinal Permeability and Cytokine Production In Vitro
Marta Calatayud, Rosa Aragao Börner, Jonas Ghyselinck, Lynn Verstrepen, Jelle De Medts, Pieter Van den Abbeele, Claire L. Boulangé, Sarah Priour, Massimo Marzorati, Sami Damak
Nutrients.2021; 13(11): 3897. CrossRef
Genome-wide functional analysis of phosphatases in the pathogenic fungus Cryptococcus neoformans
Jae-Hyung Jin, Kyung-Tae Lee, Joohyeon Hong, Dongpil Lee, Eun-Ha Jang, Jin-Young Kim, Yeonseon Lee, Seung-Heon Lee, Yee-Seul So, Kwang-Woo Jung, Dong-Gi Lee, Eunji Jeong, Minjae Lee, Yu-Byeong Jang, Yeseul Choi, Myung Ha Lee, Ji-Seok Kim, Seong-Ryong Yu,
Nature Communications.2020;[Epub] CrossRef
NEGATIVE ION MASS SPECTROMETRY FOR THE ANALYSIS OF N‐LINKED GLYCANS
David J. Harvey
Mass Spectrometry Reviews.2020; 39(5-6): 586. CrossRef
Yeast synthetic biology for designed cell factories producing secretory recombinant proteins
Eun Jung Thak, Su Jin Yoo, Hye Yun Moon, Hyun Ah Kang
FEMS Yeast Research.2020;[Epub] CrossRef
Core N -Glycan Structures Are Critical for the Pathogenicity of Cryptococcus neoformans by Modulating Host Cell Death
Eun Jung Thak, Su-Bin Lee, Shengjie Xu-Vanpala, Dong-Jik Lee, Seung-Yeon Chung, Yong-Sun Bahn, Doo-Byoung Oh, Mari L. Shinohara, Hyun Ah Kang, J. Andrew Alspaugh
mBio.2020;[Epub] CrossRef
Hydrophilic interaction chromatography for the analysis of biopharmaceutical drugs and therapeutic peptides: A review based on the separation characteristics of the hydrophilic interaction chromatography phases
Tohru Ikegami
Journal of Separation Science.2019; 42(1): 130. CrossRef
Impact of ∼omics in the detection and validation of potential anti-infective drugs
Nidia Maldonado-Carmona, Melissa Vázquez-Hernández, Osiris Jair Patiño Chávez, Stefany Daniela Rodríguez-Luna, Omar Jiménez Rodríguez, Sergio Sanchez, Corina Diana Ceapă
Current Opinion in Pharmacology.2019; 48: 1. CrossRef

Antagonistic effect of peptidoglycan of Streptococcus sanguinis on lipopolysaccharide of major periodontal pathogens: Sung-Hoon Lee; J. Microbiol. 2015;53(8):553-560. Published online July 31, 2015; DOI: https://doi.org/10.1007/s12275-015-5319-6

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Abstract

Streptococcus sanguinis is often found in subgingival biofilm including periodontopathogens, and is correlated with a delay in colonization by periodontopathogens. However, the effect of S. sanguinis on inflammation induced by periodontopathogens is poorly understood. Thus, this study investigated the effect of S. sanguinis peptidoglycan (PGN) on induction of TNF-α, IL-6, and IL-8 expression by lipopolysaccharide (LPS) of periodontal pathogens. LPS was extracted from Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Tannerella forsythia, and PGN was isolated from S. sanguinis. THP-1 cells, a monocytic cell-line, were cotreated with LPS of the periodontal pathogens and S. sanguinis PGN, and then the expression of inflammatory cytokines was analyzed by real-time RT-PCR. To analyze the underlying mechanism, the binding assay of the LPS to CD14 or LPS-binding protein (LBP) was performed in the presence or absence of the PGN after coating recombinant human CD14 and LBP on EIA plate. The PGN inhibited the binding of LPS to CD14 and LBP in a dose-dependent manner. Also, THP-1 cells were co-treated with the LPS in the presence of N-acetylmuramic acid and N-acetylglucosamine, as components of PGN, and the competition binding assay to CD14 and LBP was performed. N-acetylmuramic acid inhibited the induction of inflammatory cytokine expression by LPS and the binding of LPS to CD14 or LBP whereas Nacetylglucosamine did not show such effect. Collectively, the
results
suggest that S. sanguinis PGN inhibited the cytokine expression induced by the LPS of periodontopathogens due to the inhibition of LPS binding to LBP and CD14. N-acetylmuramic acid of PGN may play a role in inhibition of the LPS binding of periodontopathogens to CD14 and LBP.

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Inflammasome regulation by the cell surface ecto-5′-nucleotidase of the oral commensal, Streptococcus oralis
Natsuno Nakamura, Hirobumi Morisaki, Momoe Itsumi, Nobuo Okahashi, Haruka Fukamachi, Ayako Sato, Miki Kadena, Mariko Kikuchi, Shohei Matsui, Takahiro Funatsu, Hirotaka Kuwata
Biochemical and Biophysical Research Communications.2025; 744: 151206. CrossRef
New putative periodontopathogens and periodontal health‐associated species: A systematic review and meta‐analysis
Angéline Antezack, Damien Etchecopar‐Etchart, Bernard La Scola, Virginie Monnet‐Corti
Journal of Periodontal Research.2023; 58(5): 893. CrossRef
Correlation and mechanism between cardiac magnetic resonance imaging and oral streptococcus count in patients with primary microvascular angina pectoris
Qi Huang, Shi Sheng Wang, Rong Hua Luo
Medicine.2022; 101(12): e29060. CrossRef
Oral ecological environment modifications by hard-cheese: from pH to microbiome: a prospective cohort study based on 16S rRNA metabarcoding approach
Erna Cecilia Lorenzini, Barbara Lazzari, Gianluca Martino Tartaglia, Giampietro Farronato, Valentina Lanteri, Sara Botti, Filippo Biscarini, Paolo Cozzi, Alessandra Stella
Journal of Translational Medicine.2022;[Epub] CrossRef
Biofilm growth and IL-8 & TNF-α-inducing properties of Candida albicans in the presence of oral gram-positive and gram-negative bacteria
Radhika G. Bhardwaj, Arjuna Ellepolla, Hana Drobiova, Maribasappa Karched
BMC Microbiology.2020;[Epub] CrossRef
Genetics ofsanguinis-Group Streptococci in Health and Disease
Angela Nobbs, Jens Kreth, Vincent A. Fischetti, Richard P. Novick, Joseph J. Ferretti, Daniel A. Portnoy, Miriam Braunstein, Julian I. Rood
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Hao Wu, Li Xie, Min He, Ruitao Zhang, Yuan Tian, Suru Liu, Tao Gong, Fangjun Huo, Ting Yang, Qingyuan Zhang, Shujuan Guo, Weidong Tian
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Jingjing Xu, Hang Yang, Yongli Bi, Wuyou Li, Hongping Wei, Yuhong Li
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Daniela Micheline dos Santos, Emily Vivianne Freitas da Silva, Adaias Oliveira Matos, Beatriz Cristiane Zuin Monteiro, Rodrigo Antonio de Medeiros, Sandro Basso Bitencourt, Valentim Adelino Ricardo Barão, Elidiane Cipriano Rangel, Marcelo Coelho Goiato
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Research Support, Non-U.S. Gov'ts

Remodeling of the Glycosylation Pathway in the Methylotrophic Yeast Hansenula polymorpha to Produce Human Hybrid-Type N-Glycans: Seon Ah Cheon , Hyunah Kim , Doo-Byoung Oh , Ohsuk Kwon , Hyun Ah Kang; J. Microbiol. 2012;50(2):341-348. Published online April 27, 2012; DOI: https://doi.org/10.1007/s12275-012-2097-2

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Abstract: As a step forward to achieve the generation of human complex- type N-glycans in the methylotrophic yeast Hansenula polymorpha, we here report the modification of the yeast glycosylation pathway by heterologous expression of the human gene encoding β-1,2-N-acetylglucosaminyltransferase I (GnTI). For the optimal expression of human GnTI in the yeast Golgi compartment, the catalytic domain of the GnTI was fused to various N-terminal leader sequences derived from the yeast type II membrane proteins. The vectors containing GnTI fusion constructs were introduced into the H. polymorpha och1Δ single and och1Δalg3Δ double mutant strains expressing the ER-targeted Aspergillus saitoi α-1,2 mannosidase, respectively. Both of the glycoengineered Hpoch1Δ and Hpoch1ΔHpalg3Δ strains were shown to produce successfully the hybrid-type glycans with a monoantennary N-acetylglucosamine (GlcNAc1Man5GlcNAc2 and GlcNAc1Man3GlcNAc2, respectively) by N-glycan profile analysis of cell wall proteins. Furthermore, by comparative analysis of byproduct formation and the glycosylation site occupancy, we propose that the Hpoch1Δ strain would be more suitable than the Hpoch1ΔHpalg3Δ strain as a host for the production of recombinant proteins with humanized glycans.

Acinetobacter baumannii Outer Membrane Protein A Modulates the Biogenesis of Outer Membrane Vesicles: Dong Chan Moon , Chul Hee Choi , Jung Hwa Lee , Chi-Won Choi , Hye-Yeon Kim , Jeong Soon Park , Seung Il Kim , Je Chul Lee; J. Microbiol. 2012;50(1):155-160. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1589-4

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Abstract: Acinetobacter baumannii secretes outer membrane vesicles (OMVs) during both in vitro and in vivo growth, but the biogenesis mechanism by which A. baumannii produces OMVs remains undefined. Outer membrane protein A of A. baumannii (AbOmpA) is a major protein in the outer membrane and the C-terminus of AbOmpA interacts with diaminopimelate of peptidoglycan. This study investigated the role of AbOmpA in the biogenesis of A. baumannii OMVs. Quantitative and qualitative approaches were used to analyze OMV biogenesis in A. baumannii ATCC 19606T and an isogenic ΔAbOmpA mutant. OMV production was significantly increased in the ΔAbOmpA mutant compared to wild-type bacteria as demonstrated by quantitation of proteins and lipopolysaccharides (LPS) packaged in OMVs. LPS profiles prepared from OMVs from wild-type bacteria and the ΔAbOmpA mutant had identical patterns, but proteomic analysis showed different protein constituents in OMVs from wild-type bacteria compared to the ΔAbOmpA mutant. In conclusion, AbOmpA influences OMV biogenesis by controlling OMV production and protein composition.

Staphylococcal methicillin resistance expression under various growth conditions: Lee, Yoo Nik , Poo Ha Ryoung , Lee, Young Ik; J. Microbiol. 1997;35(2):103-108.

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Abstract: To improve the detection of methicillin resistant staphylococci, lowered incubation temperature (30℃) and inclusion of sodium chloride in media have been empirically recommended. However, in this study, we found that sodium chloride in Peptone-Yeast Extract-K₂HPO₄(PYK) medium decreased methicillin minimum inhibitory concentrations. Divalent cations were shown to restore the expression of staphylococcal methicillin resistance. However, when it was determined by efficiency of plating, sodium chloride increased methicillin resistance expression on agar medium in which higher divalent cations were contained in the agar medium. The decrease of minimum inhibitory concentrations at 30℃ by sodium chloride occurred in Brain Heart Infusion but did not occur in other media investigated. Interestingly, both PYK and Brain Heart Infusion media had peptone, which contain cholic acids having detergent activities. Inclusion of sodium chloride in PYK caused a higher rate of autolysis. Penicillin binding protein 2a that has a low affinity to beta-lactam antibiotics, was highly inducible in methicillin resistant Staphylococcus epidermidis strains. In this study, we found that autolysins that are activated by the sodium chloride decreased the minimum inhibitory concentration at 30℃, and peptidoglycan is weakened due to the presence of methicillin. Peptone in the media may aggravate the fragile cells. However, stabilization due to the presence of divalent cations and production of penicilin binding protein 2a increase the survival of staphylococci.

Intracellular Posttranslational Modification of Aspartyl Proteinase of Candida albicans and the Role of the Glycan Region of the Enzyme: Byoung-Kuk Na , Chul-Yong Song; J. Microbiol. 2000;38(4):218-223.

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Abstract: Using two drugs, tunicamycin and brefeldin A, which affect protein processing, we investigated the intracellular processing mechanism of secreted aspartyl proteinase 1 (SAP1) of Candida albicans. Three intracellular forms of SAP1 were detected by immunoblotting using monoclonal antibody (MAb) CAP1. Their molecular weights were approximately 40, 41 and 45 kDa, respectively. The 41 kDa protein is a glycoprotein and may be the same as the extracellular form judging by its molecular mass. The 40 kDa protein was the unglycosylated form and its molecular mass coincided with deglycosylated SAP1 and the 45 kDa protein was also the unglycosylated form. Neither the 40 and 45 kDa proteins were detected in the culture supernatant of C. albicans. These suggested that the 40 and 45 kDa proteins might be intracellular precursor forms of SAP1. These results show that SAP1 is translated as a 45 kDa precusor form in the endoplasmic reticulum and the 45 kDa precursor form undergoes proteolytic cleavage after translocation into the Golgi apparatus, generating the 40 kDa precursor form. This 40 kDa precursor is converted into a 41 kDa mature form through glycosylation in the Golgi apparatus. The mature form of the 41 kDa protein is sorted into secretory vesicles and finally released into the extracellular space through membrane fusion. When the glycan region of SAP1 was digested with N-glycosidase F, both stability and activity of the enzyme decreased. These results indicate that the glycan attached to the enzyme may, at least in part, be related to enzyme stability and activity.

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