Name: İnsan Bağırsak Mikrobiyotasında C-Glikozit Metabolik Enzimlerinin Karakterizasyonu için Yapısal Biyoloji ve Analitik Kimya Yaklaşımları (Video) | JoVE Journal
Uploaded: 2025-05-23T12:50:00.000+00:00
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Description: Keşfedin, insan bağırsak mikrobiyotasındaki C-glikozit metabolik enzimlerinin yapısal biyoloji ve analitik kimya teknikleriyle karakterizasyonu. Bu yöntem, enzim aktivitelerini ve metabolik süreçleri anlamak için güçlü bir çerçeve sunar.

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5 minutes

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<table><tbody><tr><td>2,6-dikloroindofenol (DCPIP)</td><td>Solarbio Bilim ve Teknoloji</td><td>D6350</td><td></td></tr><tr><td>2-Merkaptoetanol(&beta;-ME)</td><td>Sigma-Aldrich</td><td>08168</td><td></td></tr><tr><td>&nbsp; 5 & kez; Protein Yükleme</td><td>Boyası Sangon Biotech</td><td>C508320</td><td></td></tr><tr><td>Asetik asit</td><td>Shanghai MACKLIN</td><td>A111247</td><td></td></tr><tr><td>Asetonitril</td><td>Thermo Fisher Scientific & ? Invitrogen</td><td>A12147</td><td></td></tr><tr><td>Bruker Avance III HD 700 MHz NMR Spektrometresi</td><td>Bruker</td><td>AVANCE III HD 700MHz</td><td></td></tr><tr><td>Bruker Ultrashield 400 Plus 400MHz</td><td>Bruker</td><td>AVANCE III 400 MHz</td><td></td></tr><tr><td>Santrifüj 5424R</td><td>Eppendorf</td><td>022620601</td><td></td></tr><tr><td>Chirascan & trade; artı CD spektrometresi</td><td>Uygulamalı Fotofizik</td><td>Chirascan & ticaret; &nbsp;</td><td></td></tr><tr><td>Daidzein Shanghai</td><td>Yuanye Bio-Technology</td><td>B20227</td><td></td></tr><tr><td>Daidzin</td><td>Shanghai Yuanye Bio-Technology</td><td>B20226</td><td></td></tr><tr><td>Dithiothreitol (DTT)</td> <td>Solarbio Bilim ve Teknoloji</td><td>D8220</td><td></td></tr><tr><td>E. coli BL21 (DE3) yetkin hücreler</td><td>Thermo Fisher Scientific &trade;? Invitrogen</td><td>EC0114</td><td></td></tr><tr><td>Genistein</td><td>Shanghai Yuanye Biyo-Teknoloji</td><td>S31565</td><td></td></tr><tr><td>Genistein-8-C-glukozit</td><td>Shanghai Yuanye Biyo-Teknoloji</td><td>BP2095</td><td></td></tr><tr><td>Genistin</td><td>Shanghai Yuanye Biyo-Teknoloji</td><td>S31591</td><td></td></tr><tr><td>Glikoz</td><td>Sigma-Aldrich</td><td>G8270</td><td></td></tr><tr><td>GraphPad Prism 9</td><td>GraphPad Yazılımı</td><td>Sürüm? Prizma 9</td><td>Km ve kcat kinetik parametrelerini hesaplayın</td></tr><tr><td>HiLoad Superdex 200 pg hazırlayıcı SEC sütunları</td><td>Cytiva</td><td>28989335</td><td></td></tr><tr><td>HisTrap HP Etiket protein saflaştırma kolonları</td><td>Cytiva</td><td>17524802</td><td></td></tr><tr><td>HiTrap Q FF anyon değişim kromatografisi kolon</td><td>Cytiva</td><td>17515601</td><td></td></tr><tr><td>HKL-3000</td><td>HKL araştırma A.Ş.</td><td>&nbsp; sürüm 723</td><td></td></tr><tr><td>HPLC Sütunu TC-C18 (2), 170 & 1 & 3 mikro; m, 4.6 x 250 mm</td><td>Agilent Technologies</td><td>518925-902</td><td></td></tr><tr><td>İzopropil ve beta;-D-1-tiyokalaktopiranosit? IPTG)</td><td>Sigma-Aldrich</td><td>PHG0010</td><td></td></tr><tr><td>Laktoz</td><td>Sigma-Aldrich</td><td>17814</td><td></td></tr><tr><td>LC-20AT, yüksek performanslı sıvı kromatografisi</td><td>Shimadzu</td><td>LC20AT</td><td></td></tr><tr><td>Maltoz</td><td>Sigma-Aldrich</td><td>47288</td><td></td></tr><tr><td>Maltotriose</td><td>Sigma-Aldrich</td><td>443713</td><td></td></tr><tr><td>Manganez klorür</td><td>Şanghay Yuanye Biyo-Teknoloji</td><td>S31112</td><td></td></tr><tr><td>Metanol</td><td>Thermo Fisher Bilim;? Invitrogen</td><td>M23142</td><td></td></tr><tr><td>NAD+</td><td>Solarbio Bilim ve Teknoloji</td><td>N8110</td><td></td></tr><tr><td>Olympus SZX12 Stereo Mikroskop</td><td>OLYMPUS&nbsp;</td><td>SZX7</td><td></td></tr><tr><td>fenilmetilsülfonil florür (PMSF)</td> <td>Sigma-Aldrich</td><td>P7626 ve nbsp;</td><td></td></tr><tr><td>Phoenix</td><td>Phoenix geliştirme ekibi</td><td>1.21.2</td><td>Phoenix, Phaser yazılımı ile SAD (Tek dalga boylu Anormal Dağılım) ve MR (Moleküler Değiştirme) yöntemlerini içeren, protein kristal yapısı belirleme için kapsamlı bir yazılım paketidir.</td></tr><tr><td>Hazırlayıcı yüksek performanslı sıvı kromatografı</td><td>Sular</td><td>2545/2998</td><td></td></tr><tr><td>Protein Kristal Tarama Kiti</td><td>Hampton Research</td><td>HR2-110 / HR2-112 / HR2-144 / HR2-126 / HR2-098</td><td></td></tr><tr><td>Puerarin</td><td>Shanghai Yuanye Biyo-Teknoloji</td><td>B20446</td><td></td></tr><tr><td>Çalkalama inkübatörü</td><td>Shanghai Zhichu Instrument Co., Ltd</td><td>&nbsp; ZQZY-AF8</td><td></td></tr><tr><td>Yılan Derisi ve ticareti; Diyaliz Membranı, 10 kDa MWCO, 35 mm</td><td>Thermo Fisher Scientific&trade;? Invitrogen</td><td>88245</td><td></td></tr><tr><td>Çözünür nişasta</td><td>Sigma-Aldrich</td><td>S9765</td><td></td></tr><tr><td>Sorvall LYNX 6000 Süper Hızlı Santrifüj</td><td>Thermo Fisher Scientific &amptrade;</td><td></td><td></td>75006590</tr><tr><td>SpectraMax i3x Çok Modlu Mikroplaka Okuyucu</td><td>MOLEKÜLER CİHAZLAR</td><td>SpectraMax i3x&nbsp;</td><td></td></tr><tr><td>SpectraMax QuickDrop Mikro Hacimli Spektrofotometre Özellikleri</td><td>MOLEKÜLER CİHAZLAR</td><td>QuickDrop</td><td></td></tr><tr><td>Sükroz</td><td>Sigma-Aldrich</td><td>S5016</td><td></td></tr><tr><td>SunFire C18 OBD Hazırlık Kolon</td><td>Suları</td><td>186002741</td><td></td></tr><tr><td>Thermo Fisher Scientific Q EXACTIVE</td><td>Thermo Fisher Scientific&trade;? Invitrogen</td><td>IQLAAEGAAPFALGMBDK</td><td></td></tr><tr><td>Ubikitin benzeri proteaz 1</td><td>Thermo Fisher Scientific&trade;? Invitrogen</td><td>12588018</td><td></td></tr><tr><td>Ultra Düşük Sıcaklık Dondurucu</td><td>Thermo Fisher Scientific</td><td>905GP-ULTS</td><td></td></tr><tr><td>Vakumlu santrifüj yoğunlaştırıcı</td><td>Pekin JM Teknolojisi</td><td>CV600</td><td></td></tr><tr><td>Vitexin</td><td>Sigma-Aldrich</td><td>49513</td><td></td></tr></tbody></table>

İnsan bağırsak mikrobiyotasında c-glikozit metabolik enzimlerinin karakterizasyonu için yapısal biyoloji ve analitik kimya yaklaşımları

Structural Biology and Analytical Chemistry Approaches for Characterizing C-Glycoside Metabolic Enzymes in Human Gut Microbiota

Belirtilen prosedürleri takip ederek Şekil 1, ilk olarak DgpA, DgpB ve DgpC genlerini E. coli BL21 (DE3) hücreleri. DgpB ve DgpC kararlı bir enzim kompleksi oluşturduğundan (DgpB/C kompleksi)10,26,27,34, bu iki proteini eksprese eden hücreler, birlikte saflaştırma için birleştirildi. Hücre lizizinden sonra, süpernatant 48.0...

İnsan Bağırsak Mikrobiyotasında C-Glikozit Metabolik Enzimlerinin Karakterizasyonu için Yapısal Biyoloji ve Analitik Kimya Yaklaşımları

The gut microbiota has emerged as a key modulator of host health, particularly because of its capacity to metabolize dietary glycosides. While ...

structural-biology-analytical-chemistry-approaches-for-characterizing

Research

JoVE Journal

Biochemistry

Biyokimya

Bağışıklık, organlar ve hücreler içinde glycosphingolipid antijenlerin ekspresyonu profili içgörü kazanmak için bir spesifik ve duyarlı bir yöntem açıklanmıştır. Yöntem, kimyasal olarak sentezlenmiş bir standartlarına göre yapısal analizi için glycosphingolipid moleküllerinin adım adım parçalanma sağlayan iyon tuzak kütle spektrometri avantajına sahiptir.

Glycosphingolipid Antijenler Kütle Spektrometrik Analizi

Kaynak: Hu, Y. et.al., Endobiyotikler ile İnsan Bağırsak Mikrobiyotası Arasındaki Etkileşimlerin In Vitro Banyo Fermantasyon Sistemleri Kullanılarak Analizi. J. Vis. Exp. (2019)Bu video, insan bağırsak mikrobiyotasının ekzopolisakkaritlere karşı kısa zincirli yağ asidi üretiminin analizini göstermektedir. Bu, mikrobiyal fermantasyon ürünlerinin ayrılmasını ve gaz kromatografisi kullanılarak nicelendirilmesini vurgular. Sonuçlar, ekzopolisakkaridlerin bağırsaktaki konak-patojen dinamiklerini...

Ekzopolisakkaridler üzerindeki İnsan Bağırsağı Mikrobiyal Aktivitesinin Araştırılması

Spodoptera littoralis gut ile ilişkili aktif bakteriyel topluluk, piro sıralamadır bağlanmış kararlı-izotop-tarama (SIP) ile belirlenmiştir. Bu yöntemi kullanarak, toplum içinde aktif metabolik bakteri türlerinin belirlenmesi, yüksek çözünürlük ve hassasiyet ile yapıldı.

Kültürlü ve Gut metabolik Aktif Bakterilerin Tanımlanması<em&gt; Spodoptera littoralis</em&gt; DNA Kararlı İzotop yoluyla kullanma Probing<sup&gt; 13</sup&gt; C = Glukoz

Enzim taraması için yüksek verimli bir test tanımlanmıştır. Bu çoğullanmış kullanıma hazır tahlil kiti, önceden seçilmiş Chrömer, Polymer, Hydrogel (CPH) substratları ve kompleks I, çözünmez Chromojenik Biyokütle (ICB) substratlarından oluşur. Hedef enzimler polisakkarit parçalayıcı endo-enzimler ve proteazlardır.

Yeni çözünmez Kromojenik altyapı deneyiyle kitleri kullanılarak karbonhidrat parçalayıcı Enzimlerin yüksek verimli tarama

Burada, fizyolojik şartlarda mide bağırsak, taklit Biyoreaktörler bir dizi kullanarak gut microbiota kolon tüp bebek, kültür için bir iletişim kuralı mevcut.

Uygulama gelişmiş teknoloji insan Gut Microbiota Eğitim Kültür Vitro

Burada yüksek üretilen iş algılama ve fucosylated insan sütü oligosakkaritler (HMOs), tüm hücreli Biyoalgılayıcı kullanarak miktar açıklayın. Biz de burada, bu platform HMO üretim suşları, analizini doğru uyarlaması gürültü oranı sinyal geliştirmeye odaklanan göstermek.

Fucosylated insan süt Trisaccharides genetik olarak kullanarak Biyoteknolojik bağlamda analizini biyosensörler kodlanmış

Burada açıklanan in vitro toplu fermantasyon sistemleri kullanarak endobiyotikler ve insan bağırsak mikrobiyotları arasındaki etkileşimleri araştırmak için bir protokoldür.

In Vitro Bath Fermantasyon Sistemleri Kullanılarak Endobiyotikler ve İnsan Bağırsak Mikrobiyotları Arasındaki Etkileşimlerin Analizi

Burada açıklanan serbestçe kullanılabilir araçları kullanarak 16S rRNA metagenomik sıralama ve analiz kullanarak mikrobiyom profilleme için basitleştirilmiş bir standart işletim prosedürüdür. Bu protokol mikrobiyom alanında yeni araştırmacılar yanı sıra daha yüksek bir çözünürlükte bakteriyel profilleme elde etmek için yöntemler güncellemeleri gerektiren yardımcı olacaktır.

İki adımlı PCR ve Yeni Nesil 16S rRNA Gen Sıralaması Kullanarak Mikrobiyota Analizi

Açık arama, daha önce bilinmeyen glikan bileşimleriyle süslenmiş glikopeptidlerin tanımlanmasını sağlar. Bu makalede, Acinetobacter baumannii'yi model olarak kullanan bakteriyel örnekler için açık arama ve müteakip glikan odaklı glikopeptid aramaları yapmak için kolaylaştırılmış bir yaklaşım sunulmuştur.

Acinetobacter baumannii O-bağlantılı Glikopeptitlerlerin Tanımlanması için Açık Arama Tabanlı Yaklaşımların Uygulanması

Bu çalışma, C-glikozitleri parçalayabilen bağırsak bakterilerini verimli bir şekilde taramak ve izole etmek için bir dizi standart işletim prosedürü (SOP) oluşturur.

C-Glikozit-Parçalayan Bağırsak Bakterilerinin Taranması ve İzolasyonu

Keşfedin, insan bağırsak mikrobiyotasındaki C-glikozit metabolik enzimlerinin yapısal biyoloji ve analitik kimya teknikleriyle karakterizasyonu. Bu yöntem, enzim aktivitelerini ve metabolik süreçleri anlamak için güçlü bir çerçeve sunar.

İnsan Bağırsak Mikrobiyotasında C-Glikozit Metabolik Enzimlerinin Karakterizasyonu için Yapısal Biyoloji ve Analitik Kimya Yaklaşımları (Video) | JoVE Journal

İnsan Bağırsak Mikrobiyotasında C-Glikozit Metabolik Enzimlerinin Karakterizasyonu için Yapısal Biyoloji ve Analitik Kimya Yaklaşımları

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