Name: protein purification with sec and imac
Uploaded: 2023-04-30T06:30:54.000Z
Duration: 7 min 51 s
Channel: JoVE

K12 Standards

new-york-state-science-learning-standards

New York State Science Learning Standards

High School

Life Science

matter-and-energy-in-organisms-and-ecosystems

Matter and Energy in Organisms and Ecosystems

HS-LS1-6.

chromatography-based-biomolecule-purification-methods

Chromatography-based Biomolecule Purification Methods

クロマトグラフィーによる生体分子の浄化方法

生化学、クロマトグラフィーによる精製手法を利用して、複雑な混合物から物質を分離します。生化学者で一般的に使用される 2 つの手段は、サイズ排除クロマトグラフィーおよびアフィニ ティー ・ クロマトグラフィー。サイズ排除クロマトグラフィーでは、多孔質ビーズを充填したカラムは、サイズに基づいて混合物のコンポーネントを分離します。一方、アフィニ ティー ・ クロマトグラフィーは生体分子の特定の分離のためターゲット固有の配位子を含む液相で構成される列を使用してできます。
このビデオは、サイズ排除とアフィニ ティー ・ クロマトグラフィー、それらを支配する概念の導入として提供しています。固定化金属アフィニ ティー ・ クロマトグラフィーによるヒスチジン付けられた蛋白質の浄化のためのステップバイ ステップの手順を説明します。生物化学および生物医学研究のクロマト グラフ法の両方のためのアプリケーションがプロファイルもされます。
<div class="chapter_text" id="script_0">
「クロマトグラフィー」は複雑な混合物、生体分子の特性や活動を決定することができます前に不可欠なステップからコンポーネントを分離するために使用方法の広い範囲を指します。各クロマト グラフの技術は、サンプル マトリックスとターゲット化合物によって分離のための別のメカニズムを持っています。原則と生化学に共通の 2 つの操作に焦点を当てるこのビデオ: サイズ排除と親和性クロマトグラフィー。
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<div class="chapter_text" id="script_1">
サイズ排除クロマトグラフィーまたは SEC は、サンプル中の化合物のサイズに基づいています。多孔質材料を用いた列にサンプルを含む移動相を追加: 静止した段階。サンプルの分子は、1 の 3 つのカテゴリに分類されます。
分子が大きすぎて、毛穴を入力列を最短距離を旅行します。この「排除限界分子量"上記の分子量とのあらゆる種は同時に列を終了します。毛穴を自由に入力するには十分に小さい分子、最長保持されるため、一緒に列を終了します。完全な細孔のエントリを許可する分子量は「浸透の制限」です。
毛穴の出入りを拡散時間の様々 な量を費やすと、これらの制限の間の分子だけが、互いに区切られました。小さい分子が長時間保持される列に彼らは静止した段階のより多くの時間を過ごすためこれらの制限内で分子の大きい終了以前に対し。
1 に 2 桁全体の分子量は、これらの制限内で落ちる。心の中でこの列を選択または目的化合物の広い範囲がある場合は、シリーズで複数の列を使用できます。
今では秒の理論を見てきたを実行する方法を見てみましょう。
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<div class="chapter_text" id="script_2">
秒処理を開始するには、列を脱イオン水とクロマトグラフィー バッファーで平衡する必要があります。準備が整うと、サンプルを格納するバッファーを列に注入します。バッファーは、低流量域プッシュされます。検出器は、何が必要な試料の存在を決定する列終了を監視します。上記の排除限界分子量の大きな分子は、同時に列を終了します。小さな分数列からは、チューブで収集されます。各分画をゲル電気泳動やその他の分析技術ターゲット分子の品質をテストします。
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<div class="chapter_text" id="script_3">
今アフィニ ティー ・ クロマトグラフィーまたは AC、蛋白質を浄化するために最も効率的な方法のいずれかで見てみましょう。多くの生体分子選択的に AC を固定相にターゲット固有のリガンドを付着によって利用する特定配位子をプロパティにバインドします。
リガンドは、および残りの部分を通過するとき混合物は列に流れる、標的分子が添付します。混合物はカラムに通した後、特異性に基づいて 2 つの溶出方法の 1 つを通してターゲット分子を収集できます。
生物特異的溶出と言うことが、「通常の」や「逆の役割」があります。通常役割生物特異的溶出ターゲット生体分子と結合する付着のリガンドと競合する、エージェントが追加されます。
逆ロール生物特異的溶出、エージェントは付着のリガンドをバインドするターゲットと競合します。2 番目の溶出タイプ、無指定の溶出ソリューションの pH、イオン強度、または極性の変更ターゲット リガンドが削減されます。「タグ」を含む蛋白質を表現できるタンパク質はリガンドを固定化することができますバインドされません、場合: 短いペプチド配列リガンドを結合するように設計。
1 つの変化は金属イオンを固定化したアフィニ ティー ・ クロマトグラフィー、略して、「IMAC」ニッケルやコバルトのような付着金属のリガンドが変更された蛋白質のヒスチジン残基に結合。分子生物学的手法によるターゲット蛋白質はヒスチジン残基、ヒスチジンのイミダゾールの側鎖を介して金属にバインド polyhistidine タグと呼ばれるを繰り返し生成されます。連結された後は、蛋白質逆役割特定溶出を介して無料のイミダゾールを収集することができます、後でダウン ストリーム アプリケーションのさまざまな。
アフィニ ティー ・ クロマトグラフィーの理論を見てきた、今、研究室で IMAC 手順を見てみましょう。
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<div class="chapter_text" id="script_4">
IMAC の静止した段階は移動相と彼の付けられた蛋白質の結合をできるようにサンプルの混合物に直接追加できます。このスラリー列は、非バインドが廃棄物に滴下を化合物スラリーのままに注がれています。スラリーのコンテナーは、残留樹脂および列に追加のサンプルを収集するリンスです。
非連結の部品が自由に流れるように樹脂を攪拌します。洗浄を追加バッファーそれらを洗い流すことができます。すべての非連結コンポーネントが削除されると、廃棄物はターゲット蛋白質を収集するためのコンテナーに置き換えられます。
イミダゾールを含むバッファーが追加、攪拌、および標的分子をバインド解除する休ませます。金属、交換してタグ付きタンパク質を解放、イミダゾールにバインドします。解放されたタンパク質は、収集され、総集編をようにイミダゾール ステップが繰り返されます。さらにサンプルを浄化するには、秒は分析の前にサンプルで実行できます。
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<div class="chapter_text" id="script_5">
今では理論やこれらの 2 つの手法の手順を見てきたいくつかの生化学的なフィールドに適用する方法を見てみましょう。
蛋白質を浄化する一般的な理由は、病気での役割を研究することです。嚢胞性線維症は、嚢胞性線維症膜コンダクタンス制御するタンパク質、または CFTR の欠陥によって引き起こされます。酵母で his タグを持つタンパク質を成長後親和性とサイズ排除クロマトグラフィーその機能の研究に続いて、蛋白質の分離を許可します。
いくつかのインスタンスで polyhistidine タグの存在は、蛋白質の構造、機能に影響することを変更できます。別の一般的なタグはマルトース結合タンパク質や MBP は、アミロースを列にバインドするバインドされます。マルトースは、複雑なを解放する使用されます。MBP の切断および純粋な目的タンパク質を生成する秒を削除できます。
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<div class="chapter_text" id="script_6">
サイズ排除と親和性クロマトグラフィーでゼウスのビデオを見てきただけ。それはテクニックの理論を覆われて、一般的な手順は、行き、技術の用途のいくつかをカバーします。
見てくれてありがとう!
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In biochemistry, chromatography-based purification methods are employed to isolate compounds from a complex mixture. Two such methods used commonly by ...

「クロマトグラフィー」とは、複雑な混合物から成分を単離するために使用される幅広い方法を指し、生体分子の特性と活性を決定するための重要なステップです。各クロマトグラフィー手法には、サンプルマトリックスとターゲット化合物に応じて、分離のメカニズムが異なります。このビデオでは、生化学に共通する2つの方法、サイズ排除クロマトグラフィーとアフィニティークロマトグラフィーの原理と操作に焦点を当てます。サイズ排除クロマトグラフィー(SEC)は、サンプル中の化合物のサイズに基づいています。サンプルを含む移動相を、多孔質材料(固定相)を含むカラムに加えます。サンプル中の分子は、3つのカテゴリのうちの1つに分類されます。細孔に入るには大きすぎる分子は、カラム内を最短距離で移動します。この「排除限界」を超える分子量を持つ分子種は、同時にカラムから排出されます。細孔に自由に入るほど小さい分子は、最も長く保持され、一緒にカラムから出ます。完全な細孔の侵入を可能にする分子量が「透過限界」です。これらの限界の間の分子のみが、細孔に出入りする時間が異なるため、互いに分離されます。小さな分子は固定相でより多くの時間を過ごすため、カラム上に長く保持されますが、これらの限界内の大きな分子は早期に排出されます。1桁から2桁の分子量は、これらの制限内に収まります。この点を考慮してカラムを選択するか、目的の化合物が広範囲にある場合は、複数のカラムを直列に使用することができます。SECの理論を見てきたところで、それがどのように実行されるかを見てみましょう。SEC手順を開始するには、カラムを脱イオン水およびクロマトグラフィーバッファーで平衡化する必要があります。調製後、サンプルを含むバッファーをカラムに注入します。その後、バッファは低流量で押し出されます。検出器は、カラムから出るものをモニターして、目的の分析種の存在を判断します。分子量が排除限界を超える大きな分子は、同時にカラムから出ます。カラムからの小さな画分はチューブに集められます。各画分は、ゲル電気泳動またはその他の分析技術によって標的分子の品質について試験されます。次に、タンパク質を精製する最も効率的な方法の1つであるアフィニティークロマトグラフィーまたはACを見てみましょう。多くの生体分子は特定のリガンドに選択的に結合しますが、ACはこの特性を利用して、標的特異的なリガンドを固定相に接着させることでこの特性を利用します。混合物がカラムを流れると、標的分子が配位子に結合し、残りが流れます。混合物がカラムを通過した後、特異性に基づく2つの溶出方法のいずれかで標的分子を収集できます。生体特異的溶出は、「正常」または「逆の役割」を有すると言えます。ノーマルロールの生体特異的溶出では、付着したリガンドと競合して標的生体分子と結合する薬剤が追加されます。逆役割の生体特異的溶出では、薬剤が標的と競合して接着したリガンドに結合します。2つ目の溶出タイプである「非特異的」溶出は、溶液のpH、イオン強度、または極性を変化させることにより、ターゲットとリガンドの結合を低下させます。固定化可能なリガンドにタンパク質が結合しない場合、そのタンパク質は「タグ」(リガンドに結合するように設計された短いペプチド配列)を含んで発現させることができます。1つのバリエーションは、固定化金属イオン親和性クロマトグラフィー、略して「IMAC」で、ニッケルやコバルトなどの接着した金属配位子が修飾タンパク質のヒスチジン残基に結合します。分子生物学的手法により、ポリヒスチジンタグと呼ばれるヒスチジン残基が繰り返されることで標的タンパク質が生成され、ヒスチジン上のイミダゾール側鎖を介して金属に結合します。結合すると、タンパク質は逆役割特異的溶出を介して遊離イミダゾールで収集でき、後でさまざまなダウンストリームアプリケーションで使用できます。アフィニティークロマトグラフィーの理論を見てきたところで、研究室でのIMAC手順を見てみましょう。IMACでは、固定相を移動相とサンプルの混合物に直接添加できるため、hisタグ付きタンパク質の結合が可能になります。次に、このスラリーがカラムに注がれ、非結合化合物が廃棄物に滴下されますが、スラリーは残ります。スラリー容器をすすぎ、残留樹脂とサンプルを回収し、カラムに添加します。樹脂を攪拌して、結合されていないコンポーネントが自由に流れるようにします。ウォッシュバッファーを追加すると、それらを洗い流すことができます。結合していないすべての成分が除去されたら、廃棄物を容器に置き換えて標的タンパク質を回収します。イミダゾールを含有する緩衝液を添加し、撹拌し、休ませて標的分子の結合を解除します。イミダゾールは金属に結合し、タグ付きタンパク質を置換して放出します。遊離したタンパク質を回収し、イミダゾールのステップを繰り返して、総回収を確保します。サンプルをさらに精製するために、分析前にサンプルに対してSECを実行できます。これら2つの技術の理論と手順を見てきたので、生化学分野での応用方法をいくつか見てみましょう。タンパク質を精製する一般的な理由は、病気におけるタンパク質の役割を研究することです。嚢胞性線維症は、嚢胞性線維症の膜貫通コンダクタンス制御タンパク質(CFTR)の欠陥によって引き起こされます。酵母でhisタグを使用してタンパク質を増殖させた後、アフィニティークロマトグラフィーとサイズ排除クロマトグラフィーの両方でタンパク質を単離し、その後、その機能の研究を行います。場合によっては、ポリヒスチジンタグの存在がタンパク質の構造を変化させ、それによってその機能に影響を与える可能性があります。別の一般的なタグは、マルトース結合タンパク質またはMBPで、これはカラム内で結合したアミロースに結合します。次に、マルトースを使用して複合体を放出します。その後、MBPをSECで切断および除去して、純粋な目的のタンパク質を産生できます。JoVEのビデオ、サイズ排除クロマトグラフィーとアフィニティークロマトグラフィーをご覧になりました。それは、技術の理論をカバーし、一般的な手順を説明し、そして技術の使用のいくつかをカバーしました。ご覧いただきありがとうございます!

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protein purification with sec and imac

Protein purification with size-exclusion chromatography and affinity chromatography helps isolate biomolecules from complex mixtures. These methods are common in biochemistry because purified samples are needed before a biomolecule’s properties and activity can be studied. The video focuses on the principles behind size-exclusion chromatography, affinity chromatography, and immobilized metal affinity chromatography, or IMAC.

Size-exclusion chromatography, or SEC, separates molecules by size. A sample moves through a column packed with porous beads, which make up the stationary phase. Large molecules that cannot enter the pores travel the shortest path and leave first. Smaller molecules enter the pores, spend more time in the column, and leave later. Molecules between the exclusion limit and the permeation limit can be separated from each other, while molecules above or below those limits exit together.

The SEC procedure begins with equilibration of the column using deionized water and chromatography buffer. Sample-containing buffer is then injected, and the liquid is pushed through at a low flow rate. A detector watches what comes out of the column, and fractions are collected in tubes. These fractions can then be checked for the target molecule by gel electrophoresis or other analytical methods.

Affinity chromatography, or AC, uses the fact that many biomolecules bind selectively to certain ligands. In this method, a target-specific ligand is attached to the stationary phase. As the mixture passes through the column, the target binds to the ligand while the rest flows through. The target can then be released by biospecific elution, which may use a competing agent in normal-role or reverse-role form, or by nonspecific elution, which changes pH, ionic strength, or polarity.

If a protein does not naturally bind to an immobilized ligand, it can be engineered with a tag. A common example is a polyhistidine tag, or his-tag. In immobilized metal ion affinity chromatography, nickel or cobalt on the column binds to histidine residues through the imidazole side chain. The tagged protein is then released with free imidazole by reverse-role specific elution and can be used in downstream applications.

The IMAC procedure begins by mixing the stationary phase with the sample so the his-tagged protein can bind. The slurry is poured into the column, and unbound material drains to waste. The resin is rinsed into the column, stirred, and washed to remove leftover unbound components. After that, a container is placed to collect the target protein, and imidazole buffer is added to release it. The imidazole binds the metal, displaces the tagged protein, and the collected step may be repeated to improve recovery. For further cleanup, SEC can be run before analysis.

These purification methods are useful in biochemical and biomedical research. One example is cystic fibrosis research, where the cystic fibrosis transmembrane conductance regulator protein, or CFTR, is grown with a his-tag in yeast and then isolated with affinity and size-exclusion chromatography. In some cases, a polyhistidine tag can change protein structure and affect function. Another tag used in protein work is maltose-binding protein, or MBP, which binds amylose in a column and is released with maltose. The MBP can then be cleaved and removed with SEC to produce the desired pure protein.

Protein Purification with SEC and IMAC

dialysis: diffusion based separation

Dialysis for Buffer Exchange in Biochemistry

Dialysis is a common biochemistry technique for buffer exchange and desalting. It separates molecules by diffusion through a semipermeable membrane, which lets small molecules pass while holding larger biomolecules inside the sample. The membrane is the key part of the setup. Its pores create a molecular weight cutoff, so a membrane such as a 10 kDa membrane usually keeps molecules larger than 10 kilodaltons inside. This cutoff is not exact, because membranes have a range of pore sizes.

enzyme assays and kinetics

Measuring Enzyme Reaction Rates

Enzyme reaction rates can be measured with assays that track how substrate or product levels change over time. Enzymes are proteins, or sometimes RNAs, that speed up chemical reactions by lowering the activation energy needed to start them. They act on a specific substrate and help form a product that supports life processes. Enzymatic reactions are often described in three steps. First, the substrate binds to the enzyme active site and forms an enzyme-substrate complex. That complex can fall...

maldi-tof mass spectrometry

How MALDI-TOF Reads Biomolecules

MALDI-TOF mass spectrometry reads biomolecules by using a laser, a matrix, and a time-of-flight analyzer. MALDI, or matrix-assisted laser desorption ionization, helps fragile molecules keep their structure while they are moved into the mass analyzer. The analyzer then separates compounds based on size and charge. In MALDI, the sample is mixed into a matrix, usually an organic compound with aromatic and conjugated double bonds. A laser pulse strikes the spot, and the matrix absorbs most of the...

tandem mass spectrometry

Tandem Mass Spectrometry for Biomolecule Analysis

Tandem mass spectrometry is used to isolate a biomolecule and break it into smaller pieces so its makeup can be studied. Because biomolecules often have large, complex structures, it can be hard to identify their molecular composition and sequence from a single measurement. By linking mass spectrometers in series, the technique first selects one ion and then analyzes the fragments it produces. The process starts with an ion source that converts the sample into ions. A mass analyzer then...

protein crystallization

Protein Crystals for Structure Analysis

Protein crystals help scientists study protein structure and function. A crystal is a solid lattice made from purified protein molecules. These crystals are useful in structural biology because they reveal details that methods like mass spectrometry and SDS-PAGE cannot show on their own. Protein crystallization is paired with recombinant protein expression and x-ray diffraction to support this work. The process starts with a large amount of very pure protein. Researchers often make that...

chromatography-based biomolecule purification methods

Protein purification with size-exclusion chromatography and affinity chromatography helps isolate biomolecules from complex mixtures. These methods are common in biochemistry because purified samples are needed before a biomolecule’s properties and activity can be studied. The video focuses on the principles behind size-exclusion chromatography, affinity chromatography, and immobilized metal affinity chromatography, or IMAC. Size-exclusion chromatography, or SEC, separates molecules by size. A...

two-dimensional gel electrophoresis

Protein Mapping with 2D Gel Electrophoresis

Protein mapping with 2D gel electrophoresis separates complex protein mixtures across two steps. The method combines isoelectric focusing, or IEF, with sodium dodecyl sulfate polyacrylamide gel electrophoresis, or SDS-PAGE. Together, these methods can resolve thousands of biomolecules from a single sample by their isoelectric point and molecular weight. The first dimension is IEF. Each protein has an isoelectric point, or pI, which is the pH where its net charge is zero. Proteins are loaded...

metabolic labeling

Tracking Cell Chemistry with Metabolic Labels

Metabolic labeling tracks cell chemistry by using chemical analogs that cells take up and build into biomolecules. These labels help scientists follow biochemical changes and modifications inside cells. The labeled products can then be studied with tools such as SDS-PAGE and NMR to learn more about metabolic pathways. Several labeling strategies are shown here. Isotopic labeling uses analogs that match the natural molecule but contain uncommon isotopes, such as carbon-13 and nitrogen-15 in an...

electrophoretic mobility shift assay (emsa)

electrophoretic-mobility-shift-assay-emsa

Detecting Protein-DNA Binding by EMSA

Electrophoretic mobility shift assay, or EMSA, is a biochemical method used to detect binding between proteins and nucleic acids. It is also called a gel shift assay. The test compares how a labeled nucleic acid moves through a gel before and after it binds a protein. EMSA is used to see whether a specific protein binds to a specific nucleic acid. First, the nucleic acid is labeled to make a probe. The transcript describes radioactive phosphorus-32 as a common label. The probe is then mixed...

photometric protein determination

Measuring Protein Concentration with Absorbance

Measuring protein concentration with absorbance is a key step in many biochemical assays. Photometric methods use the fact that samples with more light-absorbing material transmit less light. Because concentration and absorption are directly related, this makes it possible to measure an unknown sample. The core idea comes from the Beer-Lambert Law. This law states that the concentration of a light-absorbing species is proportional to its absorbance. The video also introduces the Bradford assay...

density gradient ultracentrifugation

Sucrose Gradient Separation in an Ultracentrifuge

Sucrose gradient separation in an ultracentrifuge is a lab method for isolating cell parts and biomolecules. It uses differences in density to make particles sink or rise in a liquid. A high-speed ultracentrifuge speeds up this process so components can be separated without destroying them. The basic idea starts with a suspension, which is particles mixed in a liquid solvent. Particles that are denser than the solvent sediment, while less dense particles float. A rotor inside the...

co-immunoprecipitation and pull-down assays

co-immunoprecipitation-and-pull-down-assays

Protein Interaction Capture Methods in Cells

Co-immunoprecipitation and pull-down assays are laboratory methods used to capture stable protein-protein interactions. Protein-protein interactions help control many biological functions, but many of them have not yet been identified. These assays are related to immunoprecipitation, which isolates a target protein from a mixed sample using an antibody. In co-immunoprecipitation, or CoIP, an antibody binds the target protein, and that protein is already attached to another protein that the...

reconstitution of membrane proteins

Membrane Proteins in Liposome Models

Membrane proteins can be moved into liposome models so their function can be studied outside a cell. Reconstitution means restoring an isolated biomolecule to a working form. For membrane proteins, that usually means placing a purified protein into an artificial lipid membrane. Cell membranes are made mainly of phospholipids and membrane proteins. The phospholipids form a bilayer with hydrophilic phosphate heads facing water and hydrophobic fatty acid tails facing inward. Some membrane...

f&#246;rster resonance energy transfer (fret)

FRET as a Molecular Ruler in Cells

Föster resonance energy transfer, or FRET, is a way to study close-range biochemical interactions using light-emitting molecules. A donor fluorophore can pass energy to an acceptor fluorophore when their emission and absorption spectra overlap. This transfer happens only when the pair is very close together, usually within 10 nm. Because FRET depends on distance, it is often called a molecular or spectroscopic ruler. The strength of the signal is measured as FRET efficiency, which describes...

surface plasmon resonance (spr)

SPR Biosensors for Tracking Binding Kinetics

Surface plasmon resonance, or SPR, is a way to track biomolecular binding on a label-free biosensor. It is used to measure molecular affinity, kinetics, specificity, and concentration. The method works with a thin metal layer on a prism, where changes in reflected light reveal interactions in real time. SPR sensors use a thin layer of a noble metal on the face of a prism. Light is reflected from the prism-metal interface into a photodetector. At one specific angle, called the surface plasmon...

biochemistry

このコレクションでは、親和性クロマトグラフィーなどの一般的に使用される精製方法や、MALDI-TOFなどの分析方法について紹介します。さらに、共同免疫沈降法や代謝標識法など、生体分子の相互作用や機能を評価する方法も紹介します。

Explore how size-exclusion chromatography and IMAC separate proteins by size or binding, and see how tagged proteins are purified for research.

Protein Purification with SEC and IMAC | JoVE K12

Overview

Size-Exclusion Chromatography

Operation of Size-Exclusion Chromatography

Affinity Chromatography

Operation of Affinity Chromatography

Applications

Summary

What is Photosynthesis?

Photosynthesis is a multipart, biochemical process that occurs in plants as well as in some bacteria. It captures carbon dioxide and solar energy to ...

Light as Energy

The energy required to carry out photosynthesis is light&#8212; typically electromagnetic radiation from the sun. The range of all possible wavelengths is ...

Anatomy of Chloroplasts

Green algae and plants, including green stems and unripe fruit, harbor chloroplasts&mdash;the vital organelles where photosynthesis takes place. In ...

Photosystem II

The multi-protein complex photosystem II (PS II) harvests photons and transfers their energy through its bound pigments to its reaction center, and ...

Photosystem I

Although structurally similar to photosystem II (PSII), photosystem I (PSI) is has a different electron supplier and electron acceptor.
Both these ...

The Calvin Cycle

Overview

Oxygenic photosynthesis converts approximately 200 billion tons of carbon dioxide (CO2) annually to organic compounds and produces approximately ...

C4 Pathway and CAM

Most plants use the C3 pathway for carbon fixation. However, some plants, such as sugar cane, corn, and cacti that grow in hot conditions, use alternative ...

the-z-scheme-of-electron-transport-in-photosynthesis

The Z-Scheme of Electron Transport in Photosynthesis

The light reactions of photosynthesis assume a linear flow of electrons from water to NADP+. During this process, light energy drives the splitting of ...

Photosynthesis

Autotrophs

Almost all living organisms on Earth depend on photosynthesis, which is the process that converts sunlight energy into a simple sugar called ...

Plant Pigment Chromatography and Floating Leaf Discs in a Vacuum
ExpandOn the day before the experiment, fill a container with water.
Place the spinach ...

Plant Pigment Chromatography
ExpandNOTE: In this experiment you will separate pigments from spinach leaves using chromatography paper. Individual pigments ...

First Law of Thermodynamics

The First Law of Thermodynamics states that energy cannot be created or destroyed, only transformed. This can be demonstrated within a classic food web ...

Photosystems

Photosystems are multiprotein complexes that form the functional units of photosynthesis in plants, algae, and cyanobacteria. They are found embedded in ...

An Introduction to Cell Metabolism

In cells, critical molecules are either built by joining together individual units like amino acids or nucleotides, or broken down into smaller ...

Reconstitution of Membrane Proteins

Reconstitution is the process of returning an isolated biomolecule to its original form or function. This is particularly useful for studying membrane ...

Use a model to illustrate how photosynthesis transforms light energy into stored chemical energy.

HS-LS1-5.

Macromolecules

Biomolecules
Organisms contain a wide variety of organic molecules with numerous functions which depend on the chemical structures and properties of these ...

Solution Preparation
ExpandCAUTION: It is important to always use care when handling reagents. For example, iodine will stain clothes and skin so wear ...

Identifying Macromolecules with Indicator Solutions
ExpandHYPOTHESES: In this activity, the experimental hypothesis might be that the addition of an ...

Introduction to Carbohydrates

Carbohydrates, proteins, and fats are the primary macronutrients in the human diet. However, carbohydrates are the most favored source of energy in the ...

Carbohydrate Metabolism

Sugars as Energy Storage Molecules

Chemistry of Carbohydrates

Carbohydrates are an essential part of the diet in humans and animals. Grains, fruits, and vegetables are natural sources of carbohydrates that provide ...

Oligosaccharide Assembly

Protein glycosylation starts in the ER lumen and continues in the Golgi apparatus. Glycosyltransferases catalyze the addition of sugar molecules or ...

What are Lipids?

lipid-derived-compounds-in-the-human-body

Lipid-derived Compounds in the Human Body

Fats and lipids are crucial components in the human body. Some lipid-derived compounds, such as fat-soluble vitamins, eicosanoids, lipoproteins, and ...

Fats as Energy Storage Molecules

Structure of Lipids

Lipids include a diverse group of compounds that are largely nonpolar in nature. This is because they are hydrocarbons that include mostly nonpolar ...

Proteins and Amino Acids

Overview

Proteins are chains of amino acids that are connected by peptide bonds and folded into a 3-dimensional structure. The side chains of individual ...

Amino acids

Amino acids are the monomers that comprise proteins. Each amino acid has the same fundamental structure, which consists of a central carbon atom, or the ...

Peptide Bonds

A peptide bond covalently attaches amino acids through a dehydration reaction. One amino acid&#39;s carboxyl group and another amino acid&#39;s amino ...

Nucleic Acids and Nucleotides

Nucleic acids are the most important macromolecules for the continuity of life. They carry the cell&#39;s genetic blueprint and have instructions for its ...

Nucleic Acid Structure

The pentose sugar in DNA is deoxyribose, while in RNA the pentose sugar is ribose. The difference between the sugars is the presence of the hydroxyl group ...

Conditions on Early Earth

Around 4 billion years ago, oceans began to condense on earth while volcanic eruptions released nitrogen, carbon dioxide, methane, ammonia, and hydrogen ...

ATP and Macromolecule Synthesis

Biological macromolecules are organic compounds, predominantly composed of carbon atoms. The carbon atoms are covalently bonded with hydrogen, oxygen, ...

Dietary Connections

In biological systems, most metabolic pathways are interconnected. The cellular respiration processes that convert glucose to ATP&#8212;such as ...

From DNA to Protein

The flow of genetic information in cells from DNA to mRNA to protein is described by the central dogma, which states that genes specify the sequence of ...

Overview of Nitrogen Metabolism

Nitrogen is a very important element for life because it is a major constituent of proteins and nucleic acids. It is a macronutrient, and in nature, it is ...

Compounds Essential to Human Function

The human body is composed of cells that are fundamentally made up of several different molecules. These molecules are essential to carry out all ...

an-introduction-to-molecular-developmental-biology

An Introduction to Molecular Developmental Biology

Molecular signals play a major role in the complex processes occurring during embryonic development. These signals regulate activities such as cell ...

Dehydration Synthesis

Dehydration synthesis (also called a condensation reaction) is the chemical process in which two molecules covalently link together to form a new ...

an-introduction-to-saccharomyces-cerevisiae

An Introduction to Saccharomyces cerevisiae

Saccharomyces cerevisiae (commonly known as baker&rsquo;s yeast) is a single-celled eukaryote that is frequently used in scientific research.  S. ...

An Introduction to Transfection

Transfection is the process of inserting genetic material, such as DNA and double stranded RNA, into mammalian cells. The insertion of DNA into a cell ...

All living organisms on Earth are directly or indirectly dependent on photosynthesis. It is the only biological process that can capture energy from ...

an-overview-of-alkenone-biomarker-analysis-for-paleothermometry

An Overview of Alkenone Biomarker Analysis for Paleothermometry

Source: Laboratory of Jeff Salacup&#160;- University of Massachusetts Amherst
Throughout this series of videos, natural samples were extracted and ...

an-introduction-to-caenorhabditis-elegans

An Introduction to Caenorhabditis elegans

Caenorhabditis elegans is a microscopic, soil-dwelling roundworm that has been powerfully used as a model organism since the early 1970&rsquo;s.  It was ...

An Overview of Epigenetics

Since the early days of genetics research, scientists have noted certain heritable phenotypic differences that are not due to differences in the ...

An Introduction to Cell Death

Necrosis, apoptosis, and autophagic cell death are all manners in which cells can die, and these mechanisms can be induced by different stimuli, such as ...

An Overview of Gene Expression

Gene expression is the complex process where a cell uses its genetic information to make functional products. This process is regulated at multiple ...

An Introduction to Cell Division

Cell division is the process by which a parent cell divides and gives rise to two or more daughter cells. It is a means of reproduction for single-cell ...

An Overview of Genetic Analysis

An organism&#8217;s physical traits, or phenotype, are a product of its genotype, which is the combination of alleles (gene variants) inherited from its ...

an-introduction-to-cell-motility-and-migration

An Introduction to Cell Motility and Migration

Cell motility and migration play important roles in both normal biology and in disease. On one hand, migration allows cells to generate complex tissues ...

An Overview of Genetic Engineering

Genetic engineering &#8211; the process of purposefully altering an organism&#8217;s DNA &#8211; has been used to create powerful research tools and model ...

an-introduction-to-cellular-and-molecular-neuroscience

An Introduction to Cellular and Molecular Neuroscience

Cellular and molecular neuroscience is one of the newest and fastest growing subdisciplines in neuroscience. By investigating the influences of genes, ...

An Overview of Genetics and Disease

Many human diseases are associated with mutations or variations in genetic sequences. Some of these genetic variants are heritable, passed down from ...

an-introduction-to-developmental-genetics

An Introduction to Developmental Genetics

Development is the complex process through which a single-celled embryo transforms into a multicellular organism. Developmental processes are guided by ...

an-overview-of-bgdgt-biomarker-analysis-for-paleoclimatology

An Overview of bGDGT Biomarker Analysis for Paleoclimatology

Annexin V and Propidium Iodide Labeling

Staining with annexin V and propidium iodide (PI) provides researchers with a way to identify different types of cell death&#8212;either necrosis or ...

Co-Immunoprecipitation and Pull-Down Assays

Co-immunoprecipitation (CoIP) and pull-down assays are closely related methods to identify stable protein-protein interactions. These methods are related ...

Bacterial Transformation: Electroporation

The term &#x201C;transformation&#x201D; refers cellular ingestion of foreign DNA.  In nature, transformation can occur in certain types of bacteria.  In ...

Column Chromatography

Source: Laboratory of Dr. Jimmy Franco - Merrimack College
Column chromatography is one of the most useful techniques for purifying compounds. This ...

Construct and revise an explanation based on evidence for how carbon, hydrogen, and oxygen from sugar molecules may combine with other elements such as nitrogen, sulfur, and phosphorus to form amino acids and other carbon-based molecules.

Protein Purification with SEC and IMAC

Description

Transcript

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