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N-Myristoylation as a Novel Molecular Target for the Design of Chemotherapeutic Drugs
Published in Robert I. Glazer, Developments in Cancer Chemotherapy, 2019
Ronald L. Felsted, Colin Goddard, Constance J. Glover
When oligonucleotide site-directed mutagenesis was used to create mutant RSVs and Mo-MuLVs in which the amino terminal glycine codon was changed to alanine, glutamate, or aspartate codons, the resulting src and gag protein products were not myristoylated.14,28,29 In the case of the aspartate-substituted mutant protein, the initiator methionine was not removed, consistent with the known universal specificity of methionine aminopeptidase for penultimate amino acid residues.89 The initiator methionine was, however, removed from the alanine-substituted mutant proteins, but the resulting N-alanylpeptide was not myristoylated.14-29 Because amino terminal acylation was blocked by even this most conservative of amino acid substitutions, it appears that glycine is an essential prerequisite for amino terminal myristoylation. However, N-glycylproteins which are not myristoylated are known to exist.90 In addition, mutant src proteins that retain amino terminal glycine but which have other amino acid substitutions between residues 3 to 4 or 7 to 15 failed to incorporate myristate, demonstrating that the presence of the amino terminal glycine alone is not necessarily sufficient for myristoylation.28 This conclusion is also supported by the finding that certain amino acid substitutions or deletions in synthetic peptides beyond the amino terminal glycine can block or greatly reduce the rate of N-myristoyl transferase activity in vitro.70,71,86
Encephalitozoon
Published in Dongyou Liu, Handbook of Foodborne Diseases, 2018
Alexandra Valencakova, Lenka Luptakova, Monika Halanova, Olga Danisova
During a microsporidial keratoconjuctivitis and ocular microsporidiosis, a local therapeutic agent called fumagillin is recommended and applied.80 Fumagillin is an antiangiogenetic factor containing an antibiotic derived from Aspergillus fumigatus. Fumagillin and its semisynthetic analogue TNP-470 exert their effect by binding to the metalloprotease methionine aminopeptidase type 2 (MetAP2) of the pathogen and inhibiting its activity. Clinical manifestations are alleviated in less than 1 week, but for the complete elimination of the pathogen, or to prevent the recurrence of infection, a long-term application of the drug is necessary.
Fumagillin
Published in M. Lindsay Grayson, Sara E. Cosgrove, Suzanne M. Crowe, M. Lindsay Grayson, William Hope, James S. McCarthy, John Mills, Johan W. Mouton, David L. Paterson, Kucers’ The Use of Antibiotics, 2017
Current research on fumagillin is as a “backbone” for molecular development of compounds targeting signaling in cell development, differentiation, and tumorigenesis via human type 2 methionine aminopeptidase (Zhang et al., 2006). It has been suggested that fumagillin may stimulate suicidal erythrocyte death (Zbidah et al, 2012).
Therapeutic targets for the treatment of microsporidiosis in humans
Published in Expert Opinion on Therapeutic Targets, 2018
Methionine aminopeptidase (MetAPs) activity is essential for eukaryotic cell survival as the removal of the terminal methionine of a protein is often critical for its function and post-translational modification. Two major classes of MetAPs, designated type 1 and type 2 (MetAP1 and MetAP2), were originally identified as cytosolic proteins in eukaryotes [134–136]. In the genome of the cyanobacterium, Synechocystis sp., a novel MetAP3 gene was also identified [137]. MetAP2, as a member of the dimetallohydrolase family, is a cytosolic metalloenzyme that catalyzes the hydrolytic removal of N-terminal methionine residues from nascent proteins [138–140]. Important functions of this enzyme, which is found in all organisms, are its role in tissue repair and protein degradation, as well as the role it plays in angiogenesis [139,141]. The MetAP2 genes were identified from the human pathogenic microsporidia Enc. intestinalis, Enc. hellem, Enc. cuniculi, Ent. bieneusi, and A. algerae using the strategy of homology cloning by polymerase chain reaction [142–144]. Based on genome sequence data (Microsporidiadb.org), microsporidia appear to only contain MetAP2 [145]. Since both MetAP1 and MetAP2 genes exist in mammalian genomes and the functions of these two MetAP genes overlap, this makes microsporidia MetAP2 an essential gene in microsporidia and a logical target for designing therapeutic agents for microsporidiosis [144].
Development of a high-yield expression and purification system for platelet factor 4
Published in Platelets, 2018
Angela Huynh, Donald M. Arnold, Jane C. Moore, James W. Smith, John G. Kelton, Ishac Nazy
Some key disadvantages of producing eukaryotic proteins in E. coli are the limited post-translational machinery [27] and the potential for low levels of soluble and functional recombinant proteins [28]. This is demonstrated in previous methods which have been limited by low PF4 yields or low PF4 solubility. We overcame these limitations by increasing the expression levels of PF4 using a different plasmid vector designed for the overexpression of recombinant proteins in E. coli, optimizing the DNA sequence for codon bias within E. coli, and increasing solubility by adding specific detergents to the lysis process. Although glycosylation is present in other PF4 homologues, the glycosylation site on human PF4 is found in the amino-terminus which has high variability when compared to other mammalian species and is cleaved to form mature PF4 [29,30]. PF4 is approximately 7.8 kDa and undergoes minimal post-translational modifications (disulfide bonds) [31]. Therefore, the production of rhPF4 in E. coli presents a rapid and efficient method for producing PF4 for laboratory or commercial use. The primary amino acid sequence of rhPF4 produced from this purification method is identical to the human PF4 except for an additional initiating methionine at the N-terminus. E. coli normally express a methionine aminopeptidase that is able to remove the initiating methionine from expressed proteins with the subsequent amino acid being small and uncharged [32]. The methionine in the rhPF4 is not removed as a result of it being followed by a glutamic acid [33]. However, the presence of the methionine did not affect reactivity in the anti-PF4/heparin EIA or the PF4-SRA.
Obesity medications in development
Published in Expert Opinion on Investigational Drugs, 2020
Candida J. Rebello, Frank L. Greenway
Eukaryotic proteins are synthesized on the ribosome with an N-terminal methionine that in most cellular proteins is removed cotranslationally [96]. Removal of the N-terminal methionine is essential for ensuring proper functioning of these proteins many of which are important for metabolism, growth, and proliferation [97]. This processing of the N-terminal methionine is accomplished by the enzyme methionine aminopeptidase (MetAP) 2 [98]. In rodent models, MetAP2 inhibition produces reductions in body weight and body fat [99,100].