GLUT8
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GLUT8
GLUT8 also known as SLC2A8 is the eighth member of glucose transporter superfamily. It is characterized by the presence of two leucine residues in its N-terminal intracellular domain, which influences intracellular trafficking. Discovery GLUT8, originally named GLUTX1, was cloned almost simultaneously by two different groups. Tissue distribution Subcellular localization Contrary to GLUT4, GLUT8 (previously known as GLUTX1) is not insulin-sensitive. In other words, insulin does not promote GLUT8 translocation to the cell surface in neurons as well as in transfected cell lines. Where in the cell GLUT8 is localized in not yet clear. Most GLUT8 is not present at the cell surface. Some co-localization with both the endoplasmic reticulum and late endosomes/lysosomes has been published. When the N-terminal di-leucine motif is mutated into a di-alanine motif, GLUT8 is located mostly at the cell surface in Xenopus oocytes and mammalian cells such as HEK 293 cells and diffe ...
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Glucose Transporter
Glucose transporters are a wide group of membrane proteins that facilitate the transport of glucose across the plasma membrane, a process known as facilitated diffusion. Because glucose is a vital source of energy for all life, these transporters are present in all phyla. The GLUT or SLC2A family are a protein family that is found in most mammalian cells. 14 GLUTS are encoded by human genome. GLUT is a type of uniporter transporter protein. Synthesis of free glucose Most non-autotrophic cells are unable to produce free glucose because they lack expression of glucose-6-phosphatase and, thus, are involved only in glucose uptake and catabolism. Usually produced only in hepatocytes, in fasting conditions, other tissues such as the intestines, muscles, brain, and kidneys are able to produce glucose following activation of gluconeogenesis. Glucose transport in yeast In ''Saccharomyces cerevisiae'' glucose transport takes place through facilitated diffusion. The transport proteins ...
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Leucine
Leucine (symbol Leu or L) is an essential amino acid that is used in the biosynthesis of proteins. Leucine is an α-amino acid, meaning it contains an α-amino group (which is in the protonated −NH3+ form under biological conditions), an α- carboxylic acid group (which is in the deprotonated −COO− form under biological conditions), and a side chain isobutyl group, making it a non-polar aliphatic amino acid. It is essential in humans, meaning the body cannot synthesize it: it must be obtained from the diet. Human dietary sources are foods that contain protein, such as meats, dairy products, soy products, and beans and other legumes. It is encoded by the codons UUA, UUG, CUU, CUC, CUA, and CUG. Like valine and isoleucine, leucine is a branched-chain amino acid. The primary metabolic end products of leucine metabolism are acetyl-CoA and acetoacetate; consequently, it is one of the two exclusively ketogenic amino acids, with lysine being the other. It is the most import ...
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N-terminus
The N-terminus (also known as the amino-terminus, NH2-terminus, N-terminal end or amine-terminus) is the start of a protein or polypeptide, referring to the free amine group (-NH2) located at the end of a polypeptide. Within a peptide, the amine group is bonded to the carboxylic group of another amino acid, making it a chain. That leaves a free carboxylic group at one end of the peptide, called the C-terminus, and a free amine group on the other end called the N-terminus. By convention, peptide sequences are written N-terminus to C-terminus, left to right (in LTR writing systems). This correlates the translation direction to the text direction, because when a protein is translated from messenger RNA, it is created from the N-terminus to the C-terminus, as amino acids are added to the carboxyl end of the protein. Chemistry Each amino acid has an amine group and a carboxylic group. Amino acids link to one another by peptide bonds which form through a dehydration reaction that ...
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Protein Domain
In molecular biology, a protein domain is a region of a protein's polypeptide chain that is self-stabilizing and that folds independently from the rest. Each domain forms a compact folded three-dimensional structure. Many proteins consist of several domains, and a domain may appear in a variety of different proteins. Molecular evolution uses domains as building blocks and these may be recombined in different arrangements to create proteins with different functions. In general, domains vary in length from between about 50 amino acids up to 250 amino acids in length. The shortest domains, such as zinc fingers, are stabilized by metal ions or disulfide bridges. Domains often form functional units, such as the calcium-binding EF hand domain of calmodulin. Because they are independently stable, domains can be "swapped" by genetic engineering between one protein and another to make chimeric proteins. Background The concept of the domain was first proposed in 1973 by Wetlaufer aft ...
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Protein Targeting
:''This article deals with protein targeting in eukaryotes unless specified otherwise.'' Protein targeting or protein sorting is the biological mechanism by which proteins are transported to their appropriate destinations within or outside the cell. Proteins can be targeted to the inner space of an organelle, different intracellular membranes, the plasma membrane, or to the exterior of the cell via secretion. Information contained in the protein itself directs this delivery process. Correct sorting is crucial for the cell; errors or dysfunction in sorting have been linked to multiple diseases. History In 1970, Günter Blobel conducted experiments on protein translocation across membranes. Blobel, then an assistant professor at Rockefeller University, built upon the work of his colleague George Palade. Palade had previously demonstrated that non-secreted proteins were translated by free ribosomes in the cytosol, while secreted proteins (and target proteins, in general) were t ...
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Transfection
Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. It may also refer to other methods and cell types, although other terms are often preferred: "transformation" is typically used to describe non-viral DNA transfer in bacteria and non-animal eukaryotic cells, including plant cells. In animal cells, transfection is the preferred term as transformation is also used to refer to progression to a cancerous state (carcinogenesis) in these cells. Transduction is often used to describe virus-mediated gene transfer into eukaryotic cells. The word ''transfection'' is a portmanteau of ''trans-'' and ''infection''. Genetic material (such as supercoiled plasmid DNA or siRNA constructs), may be transfected. Transfection of animal cells typically involves opening transient pores or "holes" in the cell membrane to allow the uptake of material. Transfection can be carried out using calcium phosphate (i.e. tricalcium phosphate), by ele ...
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Cell Line
An immortalised cell line is a population of cells from a multicellular organism which would normally not proliferate indefinitely but, due to mutation, have evaded normal cellular senescence and instead can keep undergoing division. The cells can therefore be grown for prolonged periods ''in vitro''. The mutations required for immortality can occur naturally or be intentionally induced for experimental purposes. Immortal cell lines are a very important tool for research into the biochemistry and cell biology of multicellular organisms. Immortalised cell lines have also found uses in biotechnology. An immortalised cell line should not be confused with stem cells, which can also divide indefinitely, but form a normal part of the development of a multicellular organism. Relation to natural biology and pathology There are various immortal cell lines. Some of them are normal cell lines (e.g. derived from stem cells). Other immortalised cell lines are the ''in vitro'' equivalent ...
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Xenopus
''Xenopus'' () (Gk., ξενος, ''xenos''=strange, πους, ''pous''=foot, commonly known as the clawed frog) is a genus of highly aquatic frogs native to sub-Saharan Africa. Twenty species are currently described within it. The two best-known species of this genus are ''Xenopus laevis'' and ''Xenopus tropicalis'', which are commonly studied as model organisms for developmental biology, cell biology, toxicology, neuroscience and for modelling human disease and birth defects. The genus is also known for its polyploidy, with some species having up to 12 sets of chromosomes. Characteristics ''Xenopus laevis'' is a rather inactive creature. It is incredibly hardy and can live up to 15 years. At times the ponds that ''Xenopus laevis'' is found in dry up, compelling it, in the dry season, to burrow into the mud, leaving a tunnel for air. It may lie dormant for up to a year. If the pond dries up in the rainy season, ''Xenopus laevis'' may migrate long distances to another pond, main ...
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HEK Cell
Human embryonic kidney 293 cells, also often referred to as HEK 293, HEK-293, 293 cells, or less precisely as HEK cells, are a specific immortalised cell line derived from a spontaneously miscarried or aborted fetus or human embryonic kidney cells grown in tissue culture taken from a female fetus in 1973. HEK 293 cells have been widely used in cell biology research for many years, because of their reliable growth and propensity for transfection. They are also used by the biotechnology industry to produce therapeutic proteins and viruses for gene therapy as well as safety testing for a vast array of chemicals. 293T (or HEK 293T) is a derivative human cell line that expresses a mutant version of the SV40 large T antigen. It is very commonly used in biological research for making proteins and producing recombinant retroviruses. History HEK 293 cells were generated in 1973 by transfection of cultures of normal human embryonic kidney cells with sheared adenovirus 5 DNA in Ale ...
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PC12 Cells
PC12 is a cell line derived from a pheochromocytoma of the rat adrenal medulla, that have an embryonic origin from the neural crest that has a mixture of neuroblastic cells and eosinophilic cells. Background This cell line was first cultured by Greene and Tischler in 1976. It was developed in parallel to the adrenal chromaffin cell model because of its extreme versatility for pharmacological manipulation, ease of culture, and the large amount of information on their proliferation and differentiation. These qualities provide advantage even though they have smaller vesicles and quantal size, holding only an average of 1.9x10−19 moles of neurotransmitter released. The vesicles hold catecholamines, mostly dopamine, but also limited amount of norepinephrine, and release of these neurotransmitters give rise to spikes due to changes in current similar to chromaffin cells. PC12 cell line use has given much information to the function of proteins underlying vesicle fusion. This ...
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Acrosome
The acrosome is an organelle that develops over the anterior (front) half of the head in the spermatozoa (sperm cells) of many animals including humans. It is a cap-like structure derived from the Golgi apparatus. In placental mammals the acrosome contains degradative enzymes (including hyaluronidase and acrosin). These enzymes break down the outer membrane of the ovum, called the zona pellucida, allowing the haploid nucleus in the sperm cell to join with the haploid nucleus in the ovum. This shedding of the acrosome, or acrosome reaction, can be stimulated ''in vitro'' by substances a sperm cell may encounter naturally such as progesterone or follicular fluid, as well as the more commonly used calcium ionophore A23187. This can be done to serve as a positive control when assessing the acrosome reaction of a sperm sample by flow cytometry or fluorescence microscopy. This is usually done after staining with a fluoresceinated lectin such as FITC-PNA, FITC-PSA, FITC-ConA, or flu ...
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Spermatozoa
A spermatozoon (; also spelled spermatozoön; ; ) is a motile sperm cell, or moving form of the haploid cell that is the male gamete. A spermatozoon joins an ovum to form a zygote. (A zygote is a single cell, with a complete set of chromosomes, that normally develops into an embryo.) Sperm cells contribute approximately half of the nuclear genetic information to the diploid offspring (excluding, in most cases, mitochondrial DNA). In mammals, the sex of the offspring is determined by the sperm cell: a spermatozoon bearing an X chromosome will lead to a female (XX) offspring, while one bearing a Y chromosome will lead to a male (XY) offspring. Sperm cells were first observed in Antonie van Leeuwenhoek's laboratory in 1677. Mammalian spermatozoon structure, function, and size Humans The human sperm cell is the reproductive cell in males and will only survive in warm environments; once it leaves the male body the sperm's survival likelihood is reduced and it may die, thereby ...
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