Name: how membrane proteins move side to side
Uploaded: 2023-04-30T09:16:22.000Z
Duration: 1 min 24 s
Channel: JoVE

K12 Standards

Alabama Course of Study Science

9-12

Biology

from-molecules-to-organisms-structures-and-processes

From Molecules to Organisms: Structures and Processes

9-12.B.1.4.a

Protein Diffusion in the Membrane

생리학적 온도에서 지질 이중층은 단백질, 지질 및 기타 막 구성 요소의 확산을 허용하는 2차원 유체로 작용합니다.
측면 단백질 확산은 막 내부에서 옆으로 이동하는 것입니다.
세포외 기질은 단백질의 세포외 도메인과 상호 작용하여 이동을 방해할 수 있으며, 세포 내에서는 액틴 세포골격(actin cytoskeleton) 및 관련 막관통 단백질(transmembrane protein)이 울타리를 형성하여 코랄(corral)로 알려진 특정 구획을 생성할 수 있습니다.
이러한 세포골격 울타리는 한 우리에서 다른 우리로 단백질이 자유롭게 이동하는 것을 방해합니다. 그러나 액틴 회전율은 단백질이 한 우리에서 다른 우리로 이동할 수 있도록 울타리에 일시적인 틈을 만들 수 있습니다.
단백질의 측면 확산 속도는 광표백 또는 FRAP 후 형광 회복에 의해 결정될 수 있습니다. 이 기술에서는 일체형 멤브레인 단백질을 형광 프로브로 표지합니다.
고정된 세포에 레이저 빔이 조사되면 형광 프로브가 광표백되어 원형 비형광 영역이 생성됩니다. 손실된 형광은 표백되지 않은 단백질이 표백된 부위로 확산됨에 따라 점차 회복됩니다.

막 내 단백질 확산

단백질은 막을 가로질러 회전 확산과 측면 확산을 보여줍니다. 생쥐와 인간 세포를 융합하여 하이브리드 세포를 만드는 세포 융합 실험을 통해 단백질의 측면 확산을 확인했습니다. 인간과 생쥐 세포가 융합되었을 때 인간과 생쥐 세포의 특정 막 단백질은 각각 빨간색과 녹색 형광 마커로 표시되었습니다. 처음에는 빨간색과 녹색 형광이 세포의 각 반구에 위치했습니다. 시간이 지남에 따라 빨간색과 녹색 형광 마커로 표시된 단백질이 옆으로 움직이기 시작했고 40분 만에 전체 세포 표면에 걸쳐 완전히 혼합되었습니다. 이 실험을 통해 막 단백질이 측면 확산을 나타낼 수 있음을 확인했습니다.

광표백 후 형광 회복(FRAP) 기술을 사용하여 막 단백질의 확산 속도를 결정할 수 있습니다. 이 기술에서 표적 단백질은 단백질에 특이적인 형광단 표지 항체로 표시됩니다. 대안적으로, 유전공학을 사용하여 녹색 형광 단백질과 융합된 표적 단백질을 생산할 수도 있습니다. 레이저 빔은 무형광 표백 영역을 만드는 데 사용됩니다. 40%의 형광회복이 일어난다면, 약 40%의 막단백질이 측면확산이 가능한 것으로 간주된다. 그런 다음 표백된 영역에서 형광 회복에 필요한 시간을 사용하여 확산 속도를 계산할 수 있으며 이는 확산 계수를 결정하는 데 사용될 수 있습니다. 단백질의 위치와 구조에 따라 각 단백질은 확산계수가 다릅니다.

FRAP 연구에 따르면 모든 막 단백질이 똑같이 이동하는 것은 아니며 막 단백질의 확산 속도는 순수 지질 이중층에서 예상되는 것보다 느립니다. 막을 통한 단백질 확산 속도는 막 지질의 확산 속도보다 낮습니다. 예를 들어, 진핵 세포의 20마이크로미터 길이를 횡단하는 데 막 지질은 약 20초가 걸리는 반면 막 단백질은 최대 600~3600초가 걸릴 수 있습니다. FRAP 외에도 1) 단일 입자 추적과 같은 기타 고급 기술; 2) 막 장벽을 연구하기 위해 광학 핀셋을 사용합니다. 3) 측면 확산에서 세포질 및 외질 단백질 도메인의 역할을 확인하기 위한 유전자 변형 단백질은 막 내 단백질 확산의 다양한 측면을 탐색하는 데 사용되었습니다.

단백질은 막을 가로지르는 회전 및 측면 확산을 보여줍니다. 단백질의 측면 확산은 세포 융합을 통해 확인되었습니다 ...

Education

교육

JoVE Core

Cell Biology

세포생물학

Unit 1

제1단원

Membrane Structure and Components

막 구조 및 구성 성분

KEY TERMS AND CONCEPTS

주요 용어 및 개념

how membrane proteins move side to side

Membrane proteins can move across the cell surface by lateral diffusion, and they can also rotate in place. Scientists confirmed lateral movement with a cell fusion experiment. Mouse and human cells were fused to make hybrid cells. The membrane proteins on the two cells were tagged with red and green fluorescent markers. At first, each color stayed on its own side of the fused cell. After 40 minutes, the colors mixed across the whole cell surface, showing that the proteins had moved laterally.

FRAP, or fluorescence recovery after photobleaching, is another way to measure membrane protein diffusion. In this method, the target protein is labeled with a fluorophore-linked antibody that binds only to that protein. A genetically engineered protein fused to green fluorescent protein can also be used. A laser then creates a bleached, non-fluorescent area. If 40% of the fluorescence returns, about 40% of the proteins in that area are considered laterally diffusible.

The time needed for recovery helps scientists calculate the diffusion rate and the diffusion coefficient. Different proteins have different diffusion coefficients because their location and structure are not the same. FRAP studies also showed that membrane proteins move more slowly than expected from the pure lipid bilayer. Their diffusion is slower than membrane lipids. For example, lipids may cross the 20-micrometer length of a eukaryotic cell in about 20 seconds, while membrane proteins may take 600 to 3600 seconds.

Researchers also use other advanced methods to study membrane protein movement. These include single-particle tracking, optical tweezers to study membrane barriers, and genetically modified proteins that help show how cytoplasmic and exoplasmic domains affect lateral diffusion.

How Membrane Proteins Move Side to Side

what are membranes?

Membrane Receptors and Cell Signals

The plasma membrane sets the outer boundary of a cell and helps control how the cell interacts with its environment. It lets some substances enter, keeps others out, and releases some materials in controlled amounts. The membrane also needs to stay flexible so cells such as red and white blood cells can change shape as they move through narrow capillaries. The membrane surface also carries markers that help cells recognize one another. This recognition is important during early development,...

membrane fluidity

How Cell Membranes Stay Flexible

Cell membranes stay flexible because of the fluid mosaic model. The plasma membrane is a mosaic of phospholipids, cholesterol, proteins, and carbohydrates. These parts are separate but loosely attached, so the membrane keeps a fluid character. The membrane is still fairly rigid, but it can flow and self-seal after a very fine needle passes through it. Phospholipid tails also affect membrane fluidity. Saturated fatty acids in phospholipids have straight tails because they are packed with...

fluid mosaic model

Fluid Mosaic Model of the Membrane

The fluid mosaic model explains the structure and function of the plasma membrane. Scientists first identified the plasma membrane in the 1890s, and by 1915 they had identified its main chemical parts: lipids and proteins. Early electron micrographs showed a railroad track pattern, which helped lead to the first widely accepted membrane model in 1935.

membrane lipids

Membrane Lipids in Cells and Tissues

Membrane lipids give biological membranes their structure and help set their properties. In mammalian cells, lipids make up about 50% of the membrane on average. The amount can be much lower in the inner mitochondrial membrane or much higher in the myelin sheath around nerve cells. Several phospholipids are common in mammalian membranes. These include phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin. At physiological pH, phosphatidylserine carries a negative...

asymmetric lipid bilayer

Membrane Lipid Unevenness

Biological membranes have an uneven mix of lipids between the inner and outer layers. This membrane asymmetry gives each side a different chemical makeup and supports different cell functions. Evidence for this uneven layout came from treating an erythrocyte membrane with phospholipase. Phospholipase is an enzyme that breaks lipids into fatty acids and hydrophilic groups. It acted only on the outer layer, while the inner layer stayed intact. The treatment degraded about 80% of total membrane...

membrane asymmetry regulating transporters

membrane-asymmetry-regulating-transporters

Membrane Lipid Flipping and Transport

Membrane lipid flipping and transport help cells keep an uneven, or asymmetric, distribution of phospholipids across the membrane. This balance is controlled by flippase, floppase, and scramblase enzymes, which move lipids between the two membrane leaflets. Flippases move specific phospholipids from the outer leaflet to the inner leaflet. In eukaryotes, these enzymes are type-IV P-type ATPases, also called P4-ATPases. They are membrane pumps that use ATP and contain one transmembrane domain...

membrane carbohydrates

Membrane Sugars and Cell Identity

Membrane carbohydrates help give cells their surface identity and support plasma membrane function. The plasma membrane is a dynamic barrier made of lipids, proteins, and carbohydrates. These sugar chains have unique structural and chemical traits that help the membrane work properly for cell growth and survival. Membrane carbohydrates do not have a hydrophobic region, so they are found only on the outer surface of the cell. Sugar addition, or glycosylation, of proteins occurs in the lumens of...

membrane proteins

Facilitated Transport Across Cell Membranes

Facilitated transport uses membrane proteins to help materials cross the plasma membrane. These integral transmembrane proteins act as channels or carriers. They move selected substances through the membrane more easily than the lipid layer alone would allow. Channel proteins provide a hydrophilic, or water-friendly, path through the membrane. Their core forms a hydrated opening that lets solutes pass between the inside and outside fluids of the cell. This helps polar compounds avoid the...

lipids as anchors

Lipid Anchors in Membrane Proteins

Lipid anchors help attach proteins to the plasma membrane. In eukaryotic cells, the main lipid anchors are prenyl groups, fatty acyl groups, and glycosylphosphatidylinositol, or GPI, groups. Prenyl and fatty acyl groups anchor proteins on the cytosolic side of the membrane. GPI anchors proteins on the extracellular side. Prenylated proteins often have a CaaX motif at the carboxy terminus. In this sequence, C means cysteine, a means any aliphatic amino acid, and X means any other amino acid...

single-pass transmembrane proteins

How Membrane Proteins Span Cell Membranes

Membrane proteins help cells communicate, send signals, stick to other cells, and move molecules across the membrane. Integral membrane proteins are tightly linked to the cell membrane and carry out these jobs. Some integral membrane proteins sit only in one membrane layer. Fatty acid amide hydrolase is one example, and it is found on the cytoplasmic side of the membrane monolayer. Other integral membrane proteins are transmembrane proteins, which span across the membrane. These proteins have...

multi-pass transmembrane proteins and &#946;-barrels

multi-pass-transmembrane-proteins-and-barrels

Membrane-Spanning Helices and β-Barrels

Membrane-spanning proteins can cross the cell membrane more than once. Their transmembrane chain usually folds into either an alpha-helix or a beta-strand. Alpha-helix multi-pass proteins are common, while beta-strand proteins are found mainly in gram-negative bacteria, mitochondria, and chloroplasts. Some multi-pass alpha-helix proteins are G-protein-linked receptors, or GPCRs, and bacteriorhodopsin. GPCRs have seven transmembrane alpha-helices. Each receptor also has its own extracellular...

detergent purification of membrane proteins

detergent-purification-of-membrane-proteins

How Detergents Solubilize Membrane Proteins

Detergents help solubilize membrane proteins so they can be purified from the lipid bilayer. Their hydrophobic region can replace membrane phospholipids and keep integral membrane proteins in solution. A detergent works well only when its monomers reach the critical micelle concentration, or CMC. At that point, the molecules assemble into micelles, which are small clusters of detergent. Above the CMC, free detergent monomers stay in equilibrium with the micelles. The CMC varies with the...

protein diffusion in the membrane

Membrane proteins can move across the cell surface by lateral diffusion, and they can also rotate in place. Scientists confirmed lateral movement with a cell fusion experiment. Mouse and human cells were fused to make hybrid cells. The membrane proteins on the two cells were tagged with red and green fluorescent markers. At first, each color stayed on its own side of the fused cell. After 40 minutes, the colors mixed across the whole cell surface, showing that the proteins had moved laterally. <br...

membrane domains

Lipid Rafts and Sperm Membrane Regions

Membrane domains organize specific lipids and proteins in defined parts of a cell membrane. This arrangement supports cell signaling, adhesion, and other important cellular processes. These domains can vary in size, composition, function, and how long they last. Some membrane domains are built from proteins. The membrane can contain distinct protein groups that carry out a cell’s specific functions. A clear example is the plasma membrane of a human sperm cell, which has different proteins in...

mechanisms of membrane domain formation

How Cell Membranes Form Domains

Cell membranes form domains when lipids and proteins gather into distinct regions. These regions, or membrane domains, are shaped by different physical properties that help some molecules stay together while others move apart. Some domains form through protein-protein interactions. Others depend on specific lipids, such as sphingolipids and sterols. Large proteins, including bacteriorhodopsin, can also cluster and create visible membrane domains. Membrane proteins can be held in place by...

mechanisms of membrane-bending

How Proteins Shape Cell Membranes

Cell membranes can bend into many shapes because they are flexible and made of a fluid mosaic. That bending can be temporary, as in vesicles, or long-lasting, as in microvilli. Cells control the size, place, and time of membrane curvature. Membrane bending can start inside the membrane itself or from proteins attaching to it. Changes in lipid makeup can create curvature, and proteins can help by making the membrane asymmetric or by clustering certain lipids. The shape of integral proteins also...

key terms and concepts

Learn how membrane proteins move laterally in the membrane, how cell fusion and FRAP measure diffusion, and why proteins move slower than lipids.

How Membrane Proteins Move Side to Side | JoVE K12

cell biology

JoVE Core Cell Biology illustrates complex cellular processes in easy-to-understand yet comprehensive animated video lessons. This collection focuses on the essential components and functions of cells, including organelles and membranes, cellular transport, and cell energetics. It is complemented by scientist-in-action videos that demonstrate the application of cell biology in real-world research experiments.

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Cytoplasm

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Microtubules

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ATP Energy Storage and Release

ATP and Macromolecule Synthesis

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1. Engage in evidence-based argument to relate a cell's function to the structure, function, and diversity of its components. Examples: Muscle cells have a large number of mitochondria, which create energy for contraction in the form of ATP. Some plant cells have a large vacuole which is used for storage and transport of excess nutrients in changing environments.

9-12.B.1.1

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Types of RNA

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Eukaryotic RNA Polymerases

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Transcription Elongation Factors

pre-mrna-processing-modification-of-pre-mrna-ends

Pre-mRNA Processing: Modification of pre-mRNA Ends

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Alternative RNA Splicing

Nuclear Export of mRNA

Transfer RNA Synthesis

Ribosomal RNA Synthesis

The Nucleolus

Additional Subnuclear Structures

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Termination of Translation

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Nonsense-mediated mRNA Decay

The Eukaryotic Promoter Region

Molecular Chaperones and Protein Folding

Co-activators and Co-repressors

The Proteasome

Master Transcription Regulators

Regulated Protein Degradation

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Proteins: From Genes to Degradation

mRNA Stability and Gene Expression

Cell Specific Gene Expression

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Cis-regulatory Sequences

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Cooperative Binding of Transcription Regulators

Epigenetic Regulation

Epigenetic changes alter the physical structure of the DNA without changing the genetic sequence and often regulate whether genes are turned on or off. ...

2. Obtain and evaluate information to explain the role of DNA and RNA in transcription and translation leading to protein synthesis and cellular function.

9-12.B.1.2

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Genomic DNA in Eukaryotes

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a. Use a model to describe the structure and sequence of DNA, including nucleotide structure, base pairing, and the structure of the helix.

9-12.B.1.2.a

How Membrane Proteins Move Side to Side

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