Covalent attachment of peptides for high sensitivity solid-phase sequence analysis

Aebersold, Ruedi; Pipes, Gary D.; Wettenhall, Richard E.H.; Nika, Heinz; Hood, Leroy

doi:10.1016/0003-2697(90)90417-8

Cited by 32 publications

(11 citation statements)

References 11 publications

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“…While the present results do not represent much improvement in sensitivity when compared to conventional gas-phase sequencers, it is clear that this approach holds considerable promise for improving the sequencing sensitivity by using alternative Edman-type reagents for several reasons. One is that efficient immobilization procedures for picomole amounts of peptides or proteins on derivatized PVDF membranes 11 ) or GF disks 12 ) have already been developed. This technique will allow us to perform solid-phase sequencing on the gas-phase equipment by using FITC without further modification, with a minimal cycle time and maximal sequence yield.…”

Section: Resultsmentioning

confidence: 99%

Gas-phase Microsequencing of Peptides and Proteins with a Fluorescent Edman-type Reagent, Fluorescein Isothiocyanate

Muramoto

Nokihara

Ueda

et al. 1994

Bioscience, Biotechnology, and Biochemistry

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Section: Resultsmentioning

confidence: 99%

Gas-phase Microsequencing of Peptides and Proteins with a Fluorescent Edman-type Reagent, Fluorescein Isothiocyanate

Muramoto

Nokihara

Ueda

et al. 1994

Bioscience, Biotechnology, and Biochemistry

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“…After four washes with 9% trifluoroacetic acid-27% acetonitrile (55), the membranebound peptide was placed into an Applied Biosystems 470A sequencer. The sequencing cycles used 90% methanol-1 mM phosphate (pH 7.0) for extraction of derivatized amino acids from the membrane as described previously (1,55). The products of each cleavage reaction were collected, and 32p was detected by Cerenkov counting.…”

mentioning

confidence: 99%

Autophosphorylation of the focal adhesion kinase, pp125FAK, directs SH2-dependent binding of pp60src.

Schaller¹,

Hildebrand²,

Shannon³

et al. 1994

Mol. Cell. Biol.

1,064

946

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The phosphorylation of protein tyrosine kinases (PTKs) on tyrosine residues is a critical regulatory event that modulates catalytic activity and triggers the physical association of PTKs with Src homology 2 (SH2)-containing proteins. The integrin-linked focal adhesion kinase, pp125FAK, exhibits extracellular matrix-dependent phosphorylation on tyrosine and physically associates with two nonreceptor PTKs, pp6Orc and pp590 ', via ing sequence motif (43, 44). The specificity of such binding is dictated by the residues flanking the site of tyrosine phosphorylation as well as the structural characteristics of a particular SH2 domain (12,54). Examples drawn from growth factor receptor PTK signalling illustrate the importance of SH2-phosphotyrosine interactions in the transmission of cytoplasmic signals. For example, receptor PTK autophosphorylation can recruit SH2-containing substrates to the receptor complex, thereby facilitating their phosphorylation. In turn, the phosphorylation of certain substrates appears to be critical for the regulation of their activities (for example, the growth factordependent activation of phospholipase C&y [13,23,40]). SH2-phosphotyrosine interactions also provide a mechanism by which cytosolic enzymes whose substrates are present in cell membranes can be translocated to the proximity of their substrates (for example, phospholipase C&y and phosphatidylinositol 3-kinase, cytosolic enzymes whose substrates are membrane lipids [9,26,37], and Sos, a cytosolic protein that functions as a guanine nucleotide exchange factor for the membrane-associate protein p2lras [34]). Binding of a tyrosinephosphorylated protein to an SH2 domain-containing enzyme can lead directly to an increase in enzymatic activity. For example, binding of a tyrosine-phosphorylated peptide to the SH2 domain of the regulatory 85-kDa subunit of phosphatidylinositol 3-kinase (P13K) induces conformational changes in p85 and the concomitant increase in enzymatic activity of the holoenzyme (2,6,35,41). Finally, SH2-phosphotyrosine interactions may be critical for the negative regulation of enzymatic activity. For example, c-Src kinase activity is regulated by the binding of its SH2 domain to the C-terminal regulatory site of tyrosine phosphorylation, 8). Thus, phosphotyrosine-SH2 interactions not only mediate protein-protein complex formation but are also intimately involved in the regulation of the activity of a number of signalling molecules.The PTK pp125FAK is phosphorylated on tyrosine in response to the engagement of cell surface integrins with com-1680 on April 2, 2019 by guest

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“…Since the hexahistidine-tag was added to the C-terminals of the protein and the bands were detected with the anti-histidine antibody, this molecular sift seems to be caused by the deletion of the N-terminal region. Therefore, next we have analyzed the N-terminal amino acid sequence of the S-form using the Edman degradation method [16] and found that the S-form was processed between the N-terminal amino acids Y36 and S37 ( Figure 4B). Processing of this position is completely identical to the cleavage site of presequence of YDL178wp, which can be recognized by the mitochondrial processing protease Icp55 [17].…”

Section: Existence Of Unprocessed a Mitochondrial Enzyme: Ydl178wp Inmentioning

confidence: 99%

Existence of Unprocessed a Mitochondrial Enzyme: YDL178wp in the Membrane Fraction as an Oligomeric Formation with a Protein-Unfolding Activity

Hachiya¹

2017

MOJCSR

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Gene product of the S.cerevisiae YDL178w (YDL178wp) has been identified as three proteins; firstly, it found as an actin interacting protein 2 (AIP2) using yeast two hybrid system, then re-named D-lactate dehydrogenase 2 (Dld2) because of its enzymatic activity and mitochondrial localization using fluorescent microscopy observation with the GFP-tag at the C-terminal of the protein. Finally, we have designated it Unfoldin, since it represented robust protein-unfolding activity with the Oligomeric formation. Here we show that YDL178wp exists as the two types of molecules in yeast cells, a mature sized mitochondrial protein and a precursor sized protein in the post-mitochondrial membrane fraction. This atypical intracellular presentation with multifunctional activity exhibits gene sharing or protein moonlighting, which would be represented the pragmatism of the evolutionary process.

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Covalent attachment of peptides for high sensitivity solid-phase sequence analysis

Cited by 32 publications

References 11 publications

Gas-phase Microsequencing of Peptides and Proteins with a Fluorescent Edman-type Reagent, Fluorescein Isothiocyanate

Gas-phase Microsequencing of Peptides and Proteins with a Fluorescent Edman-type Reagent, Fluorescein Isothiocyanate

Autophosphorylation of the focal adhesion kinase, pp125FAK, directs SH2-dependent binding of pp60src.

Existence of Unprocessed a Mitochondrial Enzyme: YDL178wp in the Membrane Fraction as an Oligomeric Formation with a Protein-Unfolding Activity

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