Saturday, January 30, 2010

Amyloid P Component Peptides

Definition
Amyloid P component (AP) is a glycoprotein which is found in tissue deposits of all types of amyloid and is identical to and derived from serum amyloid P component (SAP).

Discovery
AP was not originally detected in normal, non-amyloid-containing tissues, but Schneider and Loos1 observed that fluoresceinated anti-AP antibodies stained vascular and perivascular structures in sections of a number of normal tissues and suggested that AP may be a form of type IV, basement-membrane-type collagen.

Structural Characteristics
AP is a glycoprotein composed of a pair of noncovalently bound pentameric discs with a subunit size of 23-25 kDa. Each subunit consists of 204 residues, a single disulfide bridge linking Cys 36 to Cys 95, and a carbohydrate moiety attached to Asn 32. The precursor of AP is the SAP. It shares 52% homology with the amended sequence of human C-reactive protein, an acute phase protein, and 68% homology with the Syrian hamster "female protein," another acute phase protein whose response is modulated by sex steroids. AP/SAP, C-reactive protein, and female protein belong to a family of plasma proteins called pentraxins and their considerable sequence homology is probably the result of gene duplication2.

Mode of Action
AP is apparently indistinguishable from a normal serum protein, SAP which in turn is closely related to, though distinct from, C-reactive protein, the classical acute phase reactant. Earlier report show that in mice3, though not in man, SAP behaves as an acute phase reactant and that its serum level remains persistently elevated during induction of amyloidosis by repeated injections of casein4. Human SAP undergoes calcium-dependent binding to both primary (AL) and secondary (AA) amyloid fibrils in vitro and it has been suggested that EDTA treatment may dissociate AP from a more intimate association with fibril proteins than had previously been suspected5. Furthermore, recent quantitative estimates of the relative amounts of AP and fibril protein in human amyloid indicate that, far from being a trace constituent, the ratio by weight of AP to isolated fibrils can be as high as 1:7. These observations taken together raise the possibility that SAP/AP may be involved in the pathogenesis of amyloidosis.

Functions
Association of amyloid P component with complement proteins in neurologically diseased brain tissue: In a study AP was detected in a number of lesions in human brain associated with chronic degenerative disease. These lesions included diffuse and consolidated amyloid deposits, cerebral amyloid angiopathy, neurofibrillary tangles, neuropil threads and complement activated oligodendroglia. The staining for AP generally paralleled that for complement proteins C4d and C3d, suggesting that AP may serve as an adjuvant for phagocytosis6.

Human Amyloid P Component- An Elastase Inhibitor: Amyloid deposits almost invariably contain a pentagonal-shaped protein (a so-called pentraxin), termed AP, in close opposition to the amyloid fibrils. AP is also detected alongside normal dust elastin fibres in skin and basement membrane. In a study, purified human AP was shown to inhibit the activity of porcine pancreatic elastase. The inhibition of elastolytic activity was not abolished by heating AP to 70°C. Furthermore, two other human serum proteins used as controls did not inhibit elastase activity: albumin, which has a similarly acidic pI, and C-reactive protein, which is a pentraxin, sharing 55% sequence homology with AP. Enzyme kinetic studies showed that elastase treated with AP had a slower elastolytic rate than untreated elastase. The inhibitory effect of AP was reversed by high substrate (fivefold) concentration. These observations suggest that AP may function in vivo to protect elastin and amyloid fibrils from proteolytic cleavage, indeed, this may in part account for the relative resistance of amyloid deposits to resorption and proteolysis7.

References
1. Schneider HM, Loos M (1978). Amyloid P-component--a special type of collagen? Virchows Arch. B Cell Pathol., 29(3):225-228.
2. Prelli F, Pras M, Frangione B (1985). The primary structure of human tissue amyloid P component from a patient with primary idiopathic amyloidosis. J Biol Chem., 260(24):12895-12898.
3. Pepys MB, Baltz M, Gomer K, Davies AJ, Doenhoff M. (1979). Serum amyloid P-component is an acute phase reactant in the mouse. Nature, 278(5701):259-261.
4. Baltz ML, Gomer K, Davies AJ, Evans DJ, Klaus GG, Pepys MB (1980) Differences in the acute phase responses of serum amyloid P-component (SAP) and C3 to injections of casein or bovine serum albumin in amyloid-susceptible and resistant mouse strains. Clin. exp. Immunol., 39(2):355-360.
5. Holck M, Husby G, Sletten K, Natvig JB (1979). The amyloid P-component (Protein AP): an integral part of the amyloid substance? Scand. J. Immunol., 10(1):55-60.
6. Akiyama H, Yamada T, Kawamata T, McGeer PL (1991). Association of amyloid P component with complement proteins in neurologically diseased brain tissue. Brain Res., 548(1-2):349-352.
7. Li JJ, Mc Adam KP (2006). Human Amyloid P Component: An Elastase Inhibitor. Scandinavian Journal of Immunology., 20(3):219-226.

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Procathepsin B Fragments

Definition
Procathepsin B is precursor of Cathepsin B, which is a lysosomal cysteine proteinase of the papain superfamily.

Structural Characteristics
The nucleotide sequences predict that the primary structure of preprocathepsin B contains 339 amino acids organized as follows: a 17-residue NH2-terminal prepeptide sequence followed by a 62-residue propeptide region, 254 residues in mature (single chain) cathepsin B, and a 6-residue extension at the COOH terminus. A comparison of procathepsin B sequences from three species (human, mouse, and rat) reveals that the homology between the propeptides is relatively conserved with a minimum of 68% sequence identity. In particular, two conserved sequences in the propeptide that may be functionally significant include a potential glycosylation site and the presence of a single cysteine at position 59. Comparative analysis of the three sequences also suggests that processing of procathepsin B is a multistep process, during which enzymatically active intermediate forms may be generated1.

Mode of Action
In a study, the processing mechanism of the lysosomal cysteine proteinase, cathepsin B, in mammalian cells, was investigated, thus recombinant rat and human cathepsin B precursors were expressed in Saccharomyces cerevisiae. The active-site cysteine residue was changed to serine to prevent autoprocessing. When the purified proenzymes were incubated with the soluble fraction of postnuclear organelles obtained from human hepatoma HepG2 cells, processing to a 33 kDa form corresponding to the mature endogenous single-chain enzyme was observed. Inhibitors of metallo, serine and aspartic proteinases exerted no significant effect on procathepsin B processing in vitro. Further, the processing activity was effectively blocked by cysteine proteinase inhibitors, in particular E-64 and its cathepsin-B-selective derivative CA-074. Processing positions were identified by using anti-peptide antibodies specific for epitopes in the N- and C-terminal cleavage regions. The single-chain form produced in vitro was thus shown to contain an N-terminal extension of at least four residues relative to the mature lysosomal enzyme, as well as a C-terminal extension present in the proenzyme but usually absent in fully processed cathepsin B. On expression of the wild-type proenzyme in yeast, procathepsin B undergoes autoprocessing, yielding a single-chain form of the active enzyme, which contains similar N- and C-terminal extensions. These results indicate that maturation of procathepsin B in vivo in mammalian tissues relies on the proteolytic activity of cathepsin B itself 2.

Functions
Human Procathepsin B Interacts with the Annexin II Tetramer on the Surface of Tumor Cells: To study potential roles of plasma membrane-associated extracellular cathepsin B in tumor cell invasion and metastasis, the yeast two-hybrid system was used to screen for proteins that interact with human procathepsin B. The annexin II light chain (p11), one of the two subunits of the annexin II tetramer, was one of the proteins identified. Furthermore, procathepsin B could interact with the annexin II tetramer in vivo as demonstrated by coimmunoprecipitation. Cathepsin B and the annexin II tetramer were shown by immunofluorescent staining to colocalize on the surface of human breast carcinoma and glioma cells. These results suggest that the annexin II tetramer can serve as a binding protein for procathepsin B on the surface of tumor cells, an interaction that may facilitate tumor invasion and metastasis3.

Interaction of Human Breast Fibroblasts with Collagen I Increases Secretion of Procathepsin B: Interactions of stromal and tumor cells with the extracellular matrix may regulate expression of proteases including the lysosomal proteases cathepsins B and D. A study determined that growth of breast fibroblasts on collagen I gels affected cell morphology, but not the intracellular localization of vesicles staining for cathepsin B or D. Cathepsins B and D levels (mRNA or intracellular protein) were not affected in fibroblasts growing on collagen I gels or plastic, nor was cathepsin D secreted from these cells. In contrast, protein expression and secretion of cathepsin B, primarily procathepsin B, was induced by growth on collagen I gels. The induced secretion appeared to be mediated by integrins binding to collagen I, as inhibitory antibodies against a1, a2, and ß1 integrin subunits prevented procathepsin B secretion from fibroblasts grown on collagen. Furthermore, procathepsin B secretion was induced when cells were plated on ß1 integrin antibodies. Secretion of the cysteine protease procathepsin B from breast fibroblasts may have physiological and pathological consequences, as proteases are required for normal development and for lactation of the mammary gland, yet can also initiate and accelerate the progression of breast cancer4.

References
1. Chan SJ, San Segundo B, McCormick MB, Steiner DF (1986). Nucleotide and predicted amino acid sequences of cloned human and mouse preprocathepsin B cDNAs. PNAS., 83:7721-7725.
2. Mach L, Schwihla H, Stüwe K, Rowan AD, Mort JS, Glössl J (1993). Activation of procathepsin B in human hepatoma cells: the conversion into the mature enzyme relies on the action of cathepsin B itself. Biochem. J., 293:437–442.
3. Mai J, Finley RL Jr, Waisman DM, Sloane BF (2000). Human Procathepsin B Interacts with the Annexin II Tetramer on the Surface of Tumor Cells. J. Biol. Chem., 275(17):12806-12812.
4. Koblinski JE, Dosescu J, Sameni M, Moin K, Clark K, Sloane BF (2002). Interaction of Human Breast Fibroblasts with Collagen I Increases Secretion of Procathepsin B. J Biol Chem., 277(35):32220-32227.

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Wednesday, January 27, 2010

Microbial

Definition
Microbial peptides are derived from microbes and act against host cells. Microbial peptides are usually all cyclic and they often co-occur with diketopiperazine units (DKPs) containing amino-acids found in the peptide. D-ammo-acids are usually present in some these peptides.

Discovery
This 34-residue anionic amphiphatic antimicrobial peptide named dermaseptin was purified from amphibian skin 1. Mor et al., (1994) have reported that individual dermaseptins have distinct spectra of anti-microbial activity. Some of them synergize strongly, with certain combinations showing a 100-fold increase in antibiotic activity over the activity of single peptides 2. Another microbial peptide PvMSP - 1 peptide 19 (378 - 397) fragment of malaria Plasmodium vivax (PvMSP-1) is merozoite surface antigen, T-helper epitope. This T-cell epitope is present in the merozoite surface protein-1 of PvMSP-1 3. Ovine antimicrobial anionic peptide, is the surfactant-associated ovine antimicrobial anionic peptide (AP) plays a role in protection of the respiratory tree 4, 5.

Structural Characteristics
Microbial peptides are derived wholly, or in part, by enzymatically controlled condensations and ring expansions of diketopiperazine units (DKPs) because they are virtually all cyclic and they often co-occur with DKPs containing amino-acids found in the peptide 6. Dermaseptins peptides possess a common preproregion and a variable C-terminal antimicrobial domain. Dermaseptins and related family members show extensive sequence identities with precursors of opioid peptides dermorphin, dermenkephalin, and deltorphins, as well as adenoregulin 7. Amino acid sequence of Dermaseptins-1 is ALWKTMLKKLGTMALHAGKAALGAAADTISQGTQMPF. Ovine pulmonary surfactant associated anionic peptides are smaller in size, opposite in charge, and are bactericidal in zinc saline solution 5. Hemoglobin, 3037a, Malaria FRET Substrate II sequence is DABCYL - GABA - Glu - Arg - Met - Phe - Leu - Ser - Phe - Pro – EDANS 8.

Mode of Action
Bacterial killing of ovine pulmonary surfactant appears to be due to a combination of pulmonary surfactant peptides and zinc in the serum. Ovine serum contains 15 mM zinc. The zinc is bound loosely to serum proteins, particularly albumin. The mechanisms of cell death are related to the pH of surfactant containing anionic peptide or involve a metabolic event by which the anionic peptide and zinc are taken up by the bacterial cell 5. The NH2-terminal α-helical domain 1-18 of dermaseptin has been shown to be responsible for antimicrobial activity. Dermaseptin is active against Leishmania mexicana. It stimulates microbicidal activities of polymorphonuclear leukocytes and stimulates production of reactive oxygen species, release of myeloperoxidase, and a rapid and transient elevation of cytosolic-free calcium concentration and phospholipase D activity in neutrophils 9.

Functions
Bactericidal activity: Ovine pulmonary surfactant kills several gram-positive and gram-negative bacteria in zinc saline solutions 5.

Dermaseptin: Possesses highly potent antimicrobial activity against pathogenic fungi at micromolar concentration and is not hemolytic for erythrocytes. Dermaseptin-1 has the ability to reduce the infectivity of channel catfish virus (CCV) and frog virus 3 (FV3). Dermaseptin-4 has potent spermicidal activities 9.

Malaria CSP (334 - 342): This is amino acids 334 to 342 fragment of malaria CSP derived from malaria circumsporozoite protein. It is commonly used as a control peptide for melanoma vaccine studies 10.

PvMSP - 1 peptide 19 (378 - 397): It induces lymphoproliferative responses in cells from individuals with previous P. vivax infections 3.

Hemoglobin, 3037a, Malaria FRET Substrate II: This peptide was used to characterize the molecular mechanism underlying Hb degradation by plasmepsin II (PM II). N-terminal (GABA) extension results in higher maximal velocity and dramatic concentration-dependent substrate inhibition 8.

PvMSP-1 T-cell epitopes: Linear-peptide chimeras containing the promiscuous PvMSP-1 T-cell epitopes, synthesized in tandem with the Plasmodium falciparum immunodominant circumsporozoite protein (CSP) B-cell epitope, induced high specific antibody titers, cytokine production, long-lasting immune responses, and immunoglobulin G isotype class switching in BALB/c mice. The anti-peptide antibodies generated to the CSP B-cell epitope inhibited the invasion of P. falciparum sporozoites into human hepatocytes. These data and the simplicity of design of the chimeric constructs highlight the potential of multimeric, multistage, and multispecies linear-peptide chimeras containing parasite promiscuous T-cell epitopes for malaria vaccine development 3.

References
1. Mor A, Nguyen VH, Delfour A, Migliore-Samour D, Nicolas P (1991). Isolation, amino acid sequence, and synthesis of dermaseptin, a novel antimicrobial peptide of amphibian skin. Biochemistry, 30(36): 8824-8830.
2. Mor A, Hani K, Nicolas P (1994). The vertebrate peptide antibiotics dermaseptins have overlapping structural features but target specific microorganisms. Journal of Biological Chemistry, 269(50):31635-31641.
3. Caro-Aguilar I, Rodríguez A, Calvo-Calle JM, Guzmán F, De la Vega P, Patarroyo ME, Galinski MR, Moreno A (2002). Plasmodium vivax Promiscuous T-Helper epitopes defined and evaluated as linear peptide chimera immunogens. Infect. Immun., 70:3479-3492.
4. Brogden KA, Ackermann M, Huttner KM (1998). Detection of anionic antimicrobial peptides in ovine bronchoalveolar lavage fluid and respiratory epithelium. Infection and Immunity., 66(12):5948-5954.
5. Brogden K (1996). Isolation of an ovine pulmonary surfactant-associated anionic peptide bactericidal for Pasteurella haemolytica. PNAS., 93:412-416.
6. Bycroft BW (1969). Structural Relationships in Microbial Peptides. Nature, 224:595-597.
7. Amiche M, Seon AA, Pierre TN, Nicolas P (1999). The dermaseptin precursors: a protein family with a common preproregion and a variable C-terminal antimicrobial domain. FEBS Letters., 456(3):352-356.
8. Istvan E, Goldberet DE (2005). Distal substrate interactions enhance plasmepsin activity. J. Biol. Chem., 280: 6890-6896.
9. Yamshchikov GV, Mullins DW, Chang CC, Ogino T, Thompson L, Presley J, Galavotti H, Aquila W, Deacon D, Ross W, Patterson JW, Engelhard VH, Ferrone S, Slingluff CL Jr (2005). Sequential immune escape and shifting of T cell responses in a long-term survivor of melanoma. J. Immunol., 174(11):6863-6871.

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Melanotropin-Potentiating Factor (MPF) and Analogs

Definition
Melanotropin-potentiating factor (MPF) is a C-terminal tetrapeptide of human β-lipotropin, amino acids 88 to 91. It elicits a variety of neurotrophic effects in vivo consistent with its role in neuronal regeneration.

Discovery
Carter et al., in 1979 reported MPF is the C-terminal tetrapeptide of human B-lipotropin 1. Morley et al., in 1981 described MPF analogue with high stability to proteolysis and promote Urodele Limb Regeneration 2, 3.

Structural Characteristics
The structure of MPF has been determined on the bases of chemical and physicochemical examinations which included HPLC, Edman sequence analysis combined analysis of the amino acid composition and FAB-MS analysis 4. MPF is a tetrapeptide (structure Lys-Lys-Gly-Glu) that elicits a variety of neurotrophic effects in vivo consistent with a role in neuronal regeneration 5. The peptide preferably has β-endorphin 31 (Glu or Gin) as the C-terminal amino acid residue. More preferably, the peptide includes or consists of at least β-endorphin 30-31 (Gly-Glu or Gly-Gln), even more preferably β-endorphin 29-31 (Lys-Gly-Glu or Lys-Gly-Gln), and most preferably β-endorphin 28-31 (Lys-Lys-Gly-Glu).

Mode of Action
MPF potentiates the melanotropic activity of melanocyte stimulating hormones. Although the message sequence for the melanotropic and lipolytic activity are identical for β-lipotropin, α- and β-MSH, MPF was not able to affect the lipolytic response to alpha- and beta-MSH in rabbit adipocytes. However, MPF at concentrations of 10-5 and 10-6 mol/l inhibited the lipolytic activity of β-lipotropin. The inhibition of the lipolytic response to β-lipotropin is not connected with the common lipolytic message sequence (β-LPH 47-53). Since β-lipotropin has a second lipolytic sequence in its C-terminal part this second lipolytic core of β-lipotropin might interact with MPF which has no intrinsic lipolytic activity 6.

Functions
Potentiates the melanotropic activity: MPF potentiates the interaction of β-endorphin with its brain opiate receptors. It also potentiates the melanotropic activity of melanocyte-stimulating hormones (MSH) and seems to be involved in other biological systems.

Proliferation of cultured astrocytes: MPF stimulates the proliferation of cultured astrocytes and neurite outgrowth from cultures of neocortical cholinergic and mesenchephalic dopaminergic neurons. The dose-response relationships are biphasic ("bell shaped"), maximal responses being obtained with 10-6 M concentrations of MPF. MPF and nerve growth factor act on different receptors, because their effects on cholinergic neurons are synergistic 5.

Lymphocytes stimulations: Human MPF (Lys-Lys-Gly-Glu) stimulates the proliferative response of human lymphocytes to the T-cell mitogen concanavalin A by 121-751%. Human MPF similarly stimulates rat lymphocytes, but the peak effect is seen at a 100-fold higher dose (10-6 M). Rat MPF (Lys-Lys-Gly-Gln) has a peak effect at 10-6 M with human lymphocytes, but the peak effect with rat lymphocytes is at a 1000-fold lower dose (10-9 M). Truncated forms of the MPFs (Gly-Glu, Gly-Gln, Gly, Glu, Gln) and opioid peptides (β-endorphin, [Leu] and [Met]enkephalin) show insignificant or only weak stimulatory or inhibitory effects. MPF acts via specific non-opioid receptors located on lymphocytes and that endogenously released MPF may have an important role in the functioning of the immune system 7.

Effect on corticosteroid production: Effect of MPF and β-endorphin on corticosteroid production was studied in purified isolated rat adrenal cells. Addition of MPF or β- endorphin, in doses from 5 pg to 5 μg, alone did not result in a corticosterone production. Furthermore, no effect of MPF or β-endorphin in doses from 5 pg to as high as 5 μg for both peptides upon the ACTH or α-MSH-induced corticosteroidogenesis was observed (p greater than 0.1). So both MPF and β-endorphin do not influence the steroidogenic activity in the adrenal gland. Use of these peptides for discrimination of the ACTH/α-MSH receptor interactions is suggested 8.

References
1.Carter RJ, Shuster S, Morley JS (1979). Melanotropin potentiating factor is the C-terminal tetrapeptide of human B-lipotropin. Nature, 279( 5708):74-75.
2.Morley JS (1981). MPF Analogue with High Stability to Proteolysis. Neuropeptides, 2:109-114.
3.Morley JS, Ensor DM (1989). Structural specificity of beta-endorphin c-terminal tetrapeptide (mpf) in promoting urodele limb regeneration. Life Sciences, 45:1341-1347.
4.Abiko T, Sekino H (1995). Melanotropin-potentiating factor isolated from filtrate of uremic patients suffering from melanosis and carbohydrate intolerance. Amino Acids, 9(3):229-234.
5.Owen DB, Morley JS, Ensor DM, Allen YS, Miles JB (1997). Trophic effects of melanotropin-potentiating factor (MPF) on cultures of cells of the central nervous system. Peptides, 18(7):1015-1021.
6.Richter WO, Schwandt P (1986). Melanotropin potentiating factor inhibits lipolytic activity of beta-lipotropin but not of melanocyte stimulating hormones. Neuropeptides, 7(1):73-77.
7.Owen DL, Morley JS, Ensor DM, Miles JB (1998). The C-terminal tetrapeptide of beta-endorphin (MPF) enhances lymphocyte proliferative responses. Neuropeptides, 32(2):131-139.
8.Goverde HJ, Pesman GJ, Smals AG (1988). The melanotropin potentiating factor and beta-endorphin do not modulate the alpha-melanotropin-or adrenocorticotropin-induced corticosteroidogenesis in purified isolated rat adrenal cells. Neuropeptides, 12(3):125-130.

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Melanotropin-Potentiating Factor (MPF) and Analogs

Definition
Melanotropin-potentiating factor (MPF) is a C-terminal tetrapeptide of human β-lipotropin, amino acids 88 to 91. It elicits a variety of neurotrophic effects in vivo consistent with its role in neuronal regeneration.

Discovery
Carter et al., in 1979 reported MPF is the C-terminal tetrapeptide of human B-lipotropin 1. Morley et al., in 1981 described MPF analogue with high stability to proteolysis and promote Urodele Limb Regeneration 2, 3.

Structural Characteristics
The structure of MPF has been determined on the bases of chemical and physicochemical examinations which included HPLC, Edman sequence analysis combined analysis of the amino acid composition and FAB-MS analysis 4. MPF is a tetrapeptide (structure Lys-Lys-Gly-Glu) that elicits a variety of neurotrophic effects in vivo consistent with a role in neuronal regeneration 5. The peptide preferably has β-endorphin 31 (Glu or Gin) as the C-terminal amino acid residue. More preferably, the peptide includes or consists of at least β-endorphin 30-31 (Gly-Glu or Gly-Gln), even more preferably β-endorphin 29-31 (Lys-Gly-Glu or Lys-Gly-Gln), and most preferably β-endorphin 28-31 (Lys-Lys-Gly-Glu).

Mode of Action
MPF potentiates the melanotropic activity of melanocyte stimulating hormones. Although the message sequence for the melanotropic and lipolytic activity are identical for β-lipotropin, α- and β-MSH, MPF was not able to affect the lipolytic response to alpha- and beta-MSH in rabbit adipocytes. However, MPF at concentrations of 10-5 and 10-6 mol/l inhibited the lipolytic activity of β-lipotropin. The inhibition of the lipolytic response to β-lipotropin is not connected with the common lipolytic message sequence (β-LPH 47-53). Since β-lipotropin has a second lipolytic sequence in its C-terminal part this second lipolytic core of β-lipotropin might interact with MPF which has no intrinsic lipolytic activity 6.

Functions
Potentiates the melanotropic activity: MPF potentiates the interaction of β-endorphin with its brain opiate receptors. It also potentiates the melanotropic activity of melanocyte-stimulating hormones (MSH) and seems to be involved in other biological systems.

Proliferation of cultured astrocytes: MPF stimulates the proliferation of cultured astrocytes and neurite outgrowth from cultures of neocortical cholinergic and mesenchephalic dopaminergic neurons. The dose-response relationships are biphasic ("bell shaped"), maximal responses being obtained with 10-6 M concentrations of MPF. MPF and nerve growth factor act on different receptors, because their effects on cholinergic neurons are synergistic 5.

Lymphocytes stimulations: Human MPF (Lys-Lys-Gly-Glu) stimulates the proliferative response of human lymphocytes to the T-cell mitogen concanavalin A by 121-751%. Human MPF similarly stimulates rat lymphocytes, but the peak effect is seen at a 100-fold higher dose (10-6 M). Rat MPF (Lys-Lys-Gly-Gln) has a peak effect at 10-6 M with human lymphocytes, but the peak effect with rat lymphocytes is at a 1000-fold lower dose (10-9 M). Truncated forms of the MPFs (Gly-Glu, Gly-Gln, Gly, Glu, Gln) and opioid peptides (β-endorphin, [Leu] and [Met]enkephalin) show insignificant or only weak stimulatory or inhibitory effects. MPF acts via specific non-opioid receptors located on lymphocytes and that endogenously released MPF may have an important role in the functioning of the immune system 7.

Effect on corticosteroid production: Effect of MPF and β-endorphin on corticosteroid production was studied in purified isolated rat adrenal cells. Addition of MPF or β- endorphin, in doses from 5 pg to 5 μg, alone did not result in a corticosterone production. Furthermore, no effect of MPF or β-endorphin in doses from 5 pg to as high as 5 μg for both peptides upon the ACTH or α-MSH-induced corticosteroidogenesis was observed (p greater than 0.1). So both MPF and β-endorphin do not influence the steroidogenic activity in the adrenal gland. Use of these peptides for discrimination of the ACTH/α-MSH receptor interactions is suggested 8.

References
1.Carter RJ, Shuster S, Morley JS (1979). Melanotropin potentiating factor is the C-terminal tetrapeptide of human B-lipotropin. Nature, 279( 5708):74-75.
2.Morley JS (1981). MPF Analogue with High Stability to Proteolysis. Neuropeptides, 2:109-114.
3.Morley JS, Ensor DM (1989). Structural specificity of beta-endorphin c-terminal tetrapeptide (mpf) in promoting urodele limb regeneration. Life Sciences, 45:1341-1347.
4.Abiko T, Sekino H (1995). Melanotropin-potentiating factor isolated from filtrate of uremic patients suffering from melanosis and carbohydrate intolerance. Amino Acids, 9(3):229-234.
5.Owen DB, Morley JS, Ensor DM, Allen YS, Miles JB (1997). Trophic effects of melanotropin-potentiating factor (MPF) on cultures of cells of the central nervous system. Peptides, 18(7):1015-1021.
6.Richter WO, Schwandt P (1986). Melanotropin potentiating factor inhibits lipolytic activity of beta-lipotropin but not of melanocyte stimulating hormones. Neuropeptides, 7(1):73-77.
7.Owen DL, Morley JS, Ensor DM, Miles JB (1998). The C-terminal tetrapeptide of beta-endorphin (MPF) enhances lymphocyte proliferative responses. Neuropeptides, 32(2):131-139.
8.Goverde HJ, Pesman GJ, Smals AG (1988). The melanotropin potentiating factor and beta-endorphin do not modulate the alpha-melanotropin-or adrenocorticotropin-induced corticosteroidogenesis in purified isolated rat adrenal cells. Neuropeptides, 12(3):125-130.

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Tuesday, January 26, 2010

Luteinizing Hormone-Releasing Hormones (LHRH) and Related Peptides

Definition
Luteinizing hormone-releasing hormone (LHRH), also known as Gonadotropin-Releasing Hormone (GnRH) or Luteinizing Hormone-Releasing Factor (LRF) is a hypothalamic neuropeptide which acts on the pituitary to stimulate the release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH).

Discovery
LHRH was first characterized by the groups of two Nobel Laureates Guillemin and Schally in 1977. The hypophysiotropic form of the peptide is part of a larger family of decapeptides and was the first to be discovered and characterized in mammals. The discovery of multiple LHRHs in mammalian and non-mammalian species across a wide evolutionary taxa [over 20 LHRHs identified to date] has resulted in a nomenclature based on the species each of the LHRHs was first discovered 1.

Structural Characteristics
LHRH is translated from the mRNA as a pro-hormone, which is subsequently converted to the mature decapeptide (pGlu-His- Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2) in secretory vesicles prior to its release 2. Subsequent to its secretion, LHRH may be further cleaved by soluble peptidases for the purposes of degrading, converting, or transforming 3.

Mode of Action
The effects of LHRH are mediated by high-affinity G protein-coupled LHRH-receptor (LHRH-R) on pituitary gonadotropes. In humans and most vertebrates, there are two forms of LHRHs, LHRH-I and LHRH-II that exist in the brain and in peripheral tissues. Furthermore, the processed peptide of LHRH-I, LHRH-(1–5), appears also to have biological activities that are in contrast to its parent activities. Enormous interest has been focused on LHRH-I and LHRH-II and their cognate receptors as targets for designing therapies to treat cancers of the reproductive system. LHRH-I is processed by a zinc metalloendopeptidase EC 3.4.24.15 (EP24.15) that cleaves the hormone at the fifth and sixth bond of the decapeptide (Tyr5-Gly6) to form LHRH-(1–5). Autoregulation of LHRH gene expression can also be mediated by its processed peptide, LHRH-(1–5). LHRH-(1–5) may act through one of the alternative LHRH receptors, such as the LHRH-RII. This is plausible since LHRH-(1–5) share the first 4 amino acids with at least 9 LHRH-I forms. LHRH-(1–5) have contrasting effects from its parent peptide, LHRH-I, could help explain the lack of correlation between the action of LHRH-I analogs that behave as antagonists at the pituitary level but result in agonist-like anti-proliferative effects in many cancers 4, ,5.

Functions
As growth modulatory factor: LHRH may act as a growth modulatory factor in tumors of the reproductive system. LHRH and LHRH receptors (LHRH-Rs) are expressed in human melanoma cells and LHRH-Rs are found in greater than 50% of human breast cancers 6.

Action on ovary: LHRH is released in pulses and triggers the production of gonadotropins, which stimulate the growth and release of eggs by the ovary. This fact has been best illustrated by experiments in which the actions of the decapeptide have been blocked by immunoneutralization or receptor antagonist treatment, which invariably leads to cessation or reduction of gonadotropin secretion, disruption of gonadal function, and infertility. Sterility in mutant, non-LHRH-producing mice and human infertility associated with LHRH insufficiency also provide a clear demonstration of the reproductive consequences of inappropriate or deficient LHRH neurosecretion 6. In the rat, administration of an LHRH antagonist during proestrus results in a rapid and complete inhibition of ovulation, demonstrating the importance of this neuropeptide in reproductive function.

Hypothalamic anovulation: In women, a disorder called hypothalamic anovulation occurs when the hypothalamus does not produce LHRH, which in turn results in a lack of egg production and release by the ovaries. Similarly, if the LHRH secretion pattern is altered by prolonged stressful situations, a female will show symptoms of dysmenorrhea or amenorrhea, while a male will exhibit alteration of steroidogenesis as well as spermatogenesis.

Reproductive maturation: Regulation of the synthesis and secretion of hypothalamic LHRH neurons plays a fundamental role in the process of reproductive maturation in both mammalian genders. Indeed, an increase in pulsatile LHRH release is the critical factor for the onset of puberty; hypothalamic LHRH content shows a steady increase during the first three months of life in male rats, and an increase in pulsatile LHRH release occurs at the onset of puberty in female rhesus monkeys.

References
1.Walters K, Wegorzewska IN, Chin YP, Parikh MG, Wu TJ (2008). Luteinizing Hormone-Releasing Hormone I (LHRH-I) and Its Metabolite in Peripheral Tissues. Experimental Biology and Medicine, 233 (2):123-130.
2.Wetsel WC, Mellon PL, Weiner RI, Negro-Vilar A (1991). Metabolism of pro-luteinizing hormone-releasing hormone in immortalized hypothalamic neurons. Endocrinology, 129:1584-1595.
3.Wu TJ, Mani SK, Glucksman MJ, Roberts JL (2005). Stimulation of luteinizing hormone-releasing hormone (LHRH) gene expression in GT1–7 cells by its metabolite, LHRH-(1–5). Endocrinology, 146:280–286.
4.Ramakrishnappa N, Rajamahendran R, Lin Y-M, Leung PCK (2005). GnRH in non-hypothalamic reproductive tissues. Anim Reprod Sci., 88:95-113.
5.Swanson TA, Kim SI, Myers M, Pabon A, Philibert KD, Wang M, Glucksman MJ. The role of neuropeptide processing enzymes in endocrine (prostate) cancer: EC 3.4.24.15 (EP24.15). Protein Pept Lett., 11:471-478.
6.Bajo AM, Schally AV, Halmos G, Nagy A (2003). Targeted Doxorubicin-containing Luteinizing Hormone-releasing Hormone Analogue AN-152 Inhibits the Growth of Doxorubicin-resistant MX-1 Human Breast Cancers. Clinical Cancer Research, 9: 3742-3748.

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