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Efficient biosynthesis of creatine by whole-cell catalysis from guanidinoacetic acid in Corynebacterium glutamicum

Synthetic and Systems Biotechnology 2024年第1期9卷 99-107页

作者： Chunjian Li Pengdong Sun Guoqing Wei Yuqi Zhu Jingyuan Li Yanfeng Liu Jian Chen Yang Deng College of Food Science and Engineering Qingdao Agricultural UniversityQingdao 266109China Key Laboratory of Special Food Processing(Co-construction by Ministry and Province) Ministry of Agriculture Rural AffairsQingdao Agricultural UniversityQingdao 266109China Shandong Technology Innovation Center of Special Food Qingdao 266109China Qingdao Special Food Research Institute Qingdao 266109China Qingdao Nuoan Baite Biotechnology Co. Ltd.Qingdao 266109China Science Center for Future Foods Engineering Research Center of Ministry of Education on Food Synthetic Biotechnologyand Jiangsu Province Engineering Research Center of Food Synthetic BiotechnologyJiangnan UniversityWuxi 214122China Key Laboratory of Carbohydrate Chemistry and Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxi 214122China

creatine is a naturally occurring derivative of an amino acid commonly utilized in functional foods and ***,the current industrial synthesis of creatine relies on chemical processes,which may hinder its utilization in certain ***,a biological approach was devised that employs whole-cell biocatalysis in the bacterium Corynebacterium glutamicum,which is considered safe for use in food production,to produce safe-for-consumption *** objective of this study was to identify a guanidinoacetate N-methyltransferase(GAMT)with superior catalytic activity for creatine *** employing whole-cell biocatalysis,a gamt gene from Mus caroli(Mcgamt)was cloned and expressed in *** ATCC 13032,resulting in a creatine titer of 3.37 g/***,the study employed a promoter screening strategy that utilized nine native strong promoters in *** to enhance the expression level of *** highest titer was achieved using the P1676 promoter,reaching 4.14 g/*** conditions of whole-cell biocatalysis were further optimized,resulting in a creatine titer of 5.42 g/*** is the first report of successful secretory creatine expression in ***,which provides a safer and eco-friendly approach for the industrial production of creatine.

关键词： creatine Corynebacterium glutamicum Whole-cell biocatalysis Guanidinoacetate N-Methyltransferase Food additive

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Kinetics of the Inactivation of creatine Kinase During Inhibition by Planar Anions

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Science China Chemistry 1993年第11期36卷 1315-1324页

作者：张艳陈培榕邹晓明王希成周海梦 Department of Chemistry Tsinghua University Beijing 100084 PRC Department of Biological Science and Biotechnology Tsinghua University Beijing 100084 PRC State Key Laboratory of Biomacromolecules Institute of Biophysics Academia Sinica Beijing 100101 PRC

The kinetic theory of the substrate reaction during the modification of enzyme activity previously described by Tsou has been applied to a study on the kinetics of slow irreversible inhibition of creatine kinase by planar anions. Kinetic equation of substrate reaction was derived according to the theoretical analysis and experiment data, and then was simplified. From the simplified equation for the substrate reaction in the presence of the inhibitors, the microscopic rate constants for the reaction of the inhibitors with enzyme were obtained. The mechanism of inhibition of enzyme activity was discussed.

关键词： creatine kinase inhibition activity kinetics.

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2-Chloromercuri-4-nitrophenol-modified Sites of creatine Kinase Confirmed by FPLC

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Science China Chemistry 1994年第1期37卷 50-58页

作者：张平城 National Laboratory of Biomacromolecules Institute of Biophysics Academia Sinica Beijing 100080 PRC

MNP-modified tryptic peptides of creatine kinase have been separated with two-dimensional electrophoresis, but the location of the SH groups of creatine kinase modified by MNP still seemed to be vague until recently (Laue, M. C. & Quiocho, F. A. , 1977). Two peptides have now been found to be the MNP-modified peptides with reversed-phase FPLC methods, and the amino acid compositions of these two peptides are in agreement with those of the two peptides around Cys-145 and Cys-253, respectively. These findings indicate that the two buried SH groups of creatine kinase have been modified by MNP.

关键词： creatine kinase. FPLC SH groups chemical modification mercurial.

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Hydrolysis and Cyanolysis of DTNB-Modified creatine Kinase

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Science China Chemistry 1994年第9期37卷 1082-1089页

作者：何飚王洪睿王希成周海梦 Department of Biological Science and Biotechnology Tsinghua University Beijing 100084 PRC National Laboratory of Biomacromolecules Institute of Biophysics Academia Sinica Beijing 100101 PRC

The cyanolysis of DTNB-modified creatine kinase(S,S’-di-TNB-CK)has been *** found that there exist both cyanolysis and hydrolysis at the same time under the cyanolysis condition de-scribed previously by ***-modified creatine kinase was rapidly hydrolyzed at pH 9.5 in the ab-sence of *** hydrolysis shows biphasic kinetics as seen in the semilogarithmic pseudo-first-order *** analysis shows that about one S-TNB group/mol of DTNB-modified creatine kinase was rapidly re-leased in the fast phase of the hydrolysis *** further cyanolysis of hydrolytic products showsmonophasic kinetics,and about one S-TNB group/mol of hydrolytic product was also rapidly *** results show that one of the two TNB-labeled thiol groups situated respectively at two active sites ofthe creatine kinase molecule was rapidly hydrolyzed,and the other was hydrolyzed at a very slow *** hydrolyzed products were cyanolyzed,the other residual TNB group was also *** resultssuggest that the subunits of creatine kinase are asymmetrically *** leads to the differential envi-ronments of the two thiol groups at the active sites of two *** above results also show that withthe TNB release during hydrolysis or cyanolysis,the enzymic activity was also partially recovered at the *** recovery in activity is linearly related to the extent of the regeneration of reactive thiol ***,it is suggested that the reactive thiol groups of enzyme are essential for its activity,and they arelocated in the active sites of dimeric enzyme.

关键词： creatine kinase chemical modification hydrolysis cyanolysis

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Spatial metabolomics highlights metabolic reprogramming in acute myeloid leukemia mice through creatine pathway

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Acta Pharmaceutica Sinica B 2024年第10期14卷 4461-4477页

作者： Yucheng Bao Jing Qiao Wenjie Gong Ruihong Zhang Yanting Zhou Yinyin Xie Yuan Xie Jiuming He Tong Yin Shanghai Institute of Hematology State Key Laboratory of Medical GenomicsNational Research Centre for Translational Medicine at ShanghaiRuijin Hospital affiliated to Shanghai Jiao Tong University School of MedicineShanghai 200025China College of Osteopathic Medicine Kansas City UniversityKansas CityMO 64106USA State Key Laboratory of Bioactive Substance and Function of Natural Medicines Institute of Materia MedicaChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100050China NMPA Key Laboratory for Safety Research and Evaluation of Innovative Drug Beijing 100050China

Acute myeloid leukemia(AML)is recognized as an aggressive cancer that is characterized by significant metabolic ***,we applied spatial metabolomics to achieve high-throughput,in situ identification of metabolites within the liver metastases of AML *** at metabolite and protein levels were further mapped out and validated by integrating untargeted metabolomics and *** study showed a downregulation in arginine's contribution to polyamine biosynthesis and urea cycle,coupled with an upregulation of the creatine *** upregulation of creatine synthetases Gatm and Gamt,as well as the creatine transporter Slc6a8,resulted in a marked accumulation of creatine within tumor *** process further enhances oxidative phosphorylation and glycolysis of leukemia cells,thereby boosting ATP production to foster proliferation and ***,we discovered that inhibiting Slc6a8 can counter these detrimental effects,offering a new strategy for treating AML by targeting metabolic pathways.

关键词： Spatial metabolomics Acute myeloid leukemia Metabolic reprogramming creatine Slc6a8 Oxidative phosphorylation Glycolysis Metastasis

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(Phospho)creatine:the reserve and merry-go-round of brain energetics

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Neural Regeneration Research 2023年第2期18卷 327-328页

作者： Hong-Ru Chen Ton DeGrauw Chia-Yi Kuan Department of Neurosciences University of Virginia School of MedicineCharlottesvilleVAUSA Department of Pediatrics Division of NeurologyEmory UniversityAtlantaGAUSA

creatine transporter(CrT)-deficiency,the most common form of the cerebral creatine deficiency syndromes,causes cognition impairments and severe reduction of the brain creatine(Cr)and phosphocreatine(PCr)levels,and responds poorly to oral Cr supplement as a treatment *** ca uses of cognitive impairments in CrT-deficient children remain *** recently use gene-targeting to create a mouse model of CrT-deficiency to assess the impacts of Cr/PCr deficiency on brain energetics and stress-adaptation responses(Chen et al.,2021).

关键词： impairment cerebral creatine

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Dietary glycine supplementation enhances syntheses of creatine and glutathione by tissues of hybrid striped bass(Morone saxatilis ♀ × Morone chrysops ♂) fed soybean meal-based diets

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Journal of Animal Science and Biotechnology 2024年第5期15卷 1996-2011页

作者： Wenliang He Xinyu Li Guoyao Wu Department of Animal Science Texas A&M UniversityCollege StationTX 77843USA

Background We recently reported that supplementing glycine to soybean meal-based diets is necessary for the optimum growth of 5-to 40-g(Phase-I)and 110-to 240-g(Phase-II)hybrid striped bass(HSB),as well as their intestinal *** glycine serves as an essential substrate for syntheses of creatine and glutathione(GSH)in mammals(e.g.,pigs),little is known about these metabolic pathways or their nutritional regulation in *** study tested the hypothesis that glycine supplementation enhances the activities of creatine-and GSH-forming enzymes as well as creatine and GSH availabilities in tissues of hybrid striped bass(HSB;Morone saxatilis♀×Morone chrysops♂).Methods Phase-I and Phase-II HSB were fed a soybean meal-based diet supplemented with 0%,1%,or 2%glycine for 8 *** the end of the 56-d feeding,tissues(liver,intestine,skeletal muscle,kidneys,and pancreas)were collected for biochemical *** In contrast to terrestrial mammals and birds,creatine synthesis occurred primarily in skeletal muscle from all *** liver was most active in GSH synthesis among the HSB tissues *** Phase-I HSB,supplementation with 1%or 2%glycine increased(Pcreatine(15%–19%)and hepatic GSH(8%–11%),while reducing(P0.05)in these variables between the 1%and 2%glycine *** Phase-II HSB,supplementation with 1%and 2%glycine increased(Pcreatine and GSH in the muscle(15%–27%)and liver(11%–20%)in a dose-dependent manner,with reduced ratios of hepatic GSSG/GSH in the 1%or 2%glycine *** all HSB,supplementation with 1%and 2%glycine dose-dependently increased(P<0.05)activities of intramuscular arginine:glycine amidinotransferase(22%–41%)and hepaticγ-glutamylcysteine synthetase(17%–37%),with elevated activities of intramuscular guanidinoacetate methyltransferase and hepatic GSH synthetase a

关键词： Amino acids creatine Fish Glutathione Glycine Nutrition

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Oxidized Form of creatine Kinase

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Science China Chemistry 1994年第8期37卷 964-974页

作者：王希成王帆邹晓明周海梦 Department of Biological Science and Biotechnology Tsinghua UniversityBeijing 100084PRC Permanent address:Integrated Program in CMBS College of Physicians and Surgeons of Columbia UniversityNew YorkUSA Permanent address:Department of Biochemstry and Molecular Biophysics Columbia UniversityNew YorkUSA

The purified rabbit muscle creatine kinase (R-CK) was previously considered homogeneousand without disulfide *** the method of NR/R two-dimensional diagonal SDS-PAGE,two forms of R-CK,designated respectively "oxidized form" of creatine kinase which contained intrachain disulfide bondsand "reduced form" of creatine kinase which did not have any —S—S— bridges,were for the first time *** were found to be the same in amino acid composition,in subunit molecular Weight and in isoelec-tric point,and were almost identical in enzyme *** it is hard to isolate one from the other bycommon biochemical *** extensive studies show that the oxidized form of CK also contains a pair of reactive thiol groupswhich are essential to the enzyme activity,and it has one intrachain disulfide bond per *** the nativestate,this —S—S— bond cannot be reduced by DTT,but by treating the reduced form of CK with some ox-idants,these —S—S— bonds can be formed in *** it is presumed that the disulfide bonds are cross-linked through the oxidization of two shallowly buried —SH groups.

关键词： creatine klnase oxidized form disulfide bond thiol group

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Refolding of OPTA-labeled creatine kinase denatured by guanidinium chloride

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Chinese Science Bulletin 1995年第19期40卷 1642-1646页

作者：汤震宇周海梦 Department of Biological Scienoe and Biotechnology Tsinghua University Beijing 100084 China State Key Laboratory of Macromolecules Institute of Biophysics Chinese Academy of Sciences Beijing 100101 China

Although refolding of fully denatured protein molecules in vitro is not a valid model of folding of the nascent peptide chain in a biologically active protein in vivo, it can provide some useful information for protein folding. Therefore, refolding of denatured proteins has been extensively studied in recent years. It has been previously reported that during the refolding of guanidine or urea denatured creatine kinase, the enzymatic activity and the na-

关键词： creatine kinase denaturation chemical modification refolding.

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creatine,CARBS,AND FLUIDS:HOW IMPORTANT IN SOCCER NUTRITION?

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中国运动医学杂志 2005年第4期24卷 493-500页

作者： Donald T.Kirkendall Sports Medicine Committee US Soccer Federation

KEY POINTS Because so much of the running in soccer is at less than maximal sprinting speed, creatine supplementation likely provides no benefit to match *** evidence proves that a diet rich in carbohydrates can fill muscles with glycogen, and glycogen is critical to optimal performance in soccer. Soccer players’ diets, especially in the days before hard training or competition, should include 8-10 grams of carbohydrate per kilogram of body weight (3.5-4.5 g/lb). Cereals, fruits, vegetables, breads, and pastas are good sources ofcarbohydrates. Refueling of muscle with carbohydrates should begin as soon as possible following a match or a strenuous training session. Inadequate replacement of fluids lost in sweat can lead to poor soccer performance and heat illness. Players should aim to drink enough during training sessions and matches so that their body weights after play are within about 1 kg (2.2 lb) of their starting weights.  For a light workout or an easy match, especially when the weather is cool, water can be an adequate fluid replacement, if enough is ingested. But when play is strenuous and the weather is hot, carbohydrate-electrolyte sports drinks do a better job of maintaining body fluids.

关键词： creatine CARBS FLUID

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