scholarly journals Specific cross-linking of the proline isomerase cyclophilin to a non-proline-containing peptide.

1993 ◽  
Vol 4 (2) ◽  
pp. 223-232 ◽  
Author(s):  
J A McNew ◽  
K Sykes ◽  
J M Goodman
Keyword(s):  
Amino Acids ◽  
Protein A ◽  
Protein Sequencing ◽  
Cross Linking ◽  
Candida Boidinii ◽  
Affinity Ligand ◽  
Peroxisomal Targeting ◽  
The Cross ◽  
Targeting Peptide ◽  
Carboxy Terminal

A peptide corresponding to an efficient peroxisomal targeting sequence, the carboxy terminal 12 amino acids of PMP20 from Candida boidinii, was employed as an affinity ligand to search for a peroxisomal targeting receptor. Two proteins from yeast extracts with apparent molecular masses of 20 and 80 kDa were detected by chemical cross-linking to radioiodinated peptide. Both proteins were present in cytosolic supernatants. The 20-kDa species did not cross-link to a control peptide with reversed sequence, whereas the 80-kDa protein cross-linked to both peptides. The cross-linking assay was used to purify the 20-kDa protein from Saccharomyces cerevisiae. Partial protein sequencing identified this protein as cyclophilin, the product of the CYP1 gene. This protein, a peptidyl-prolyl cis-trans isomerase, is the yeast homologue of the protein that mediates the immunosuppressant effects of the drug cyclosporin A (CsA). Cross-linking of peptide to cyclophilin was inhibited by CsA. The cross-linking of cyclophilin to the PMP20-derived peptide was unanticipated because the peptide contains no prolines. The CYP1-encoded protein was not required to target proteins to peroxisomes because this organelle appeared to be assembled normally in a CYP1-disrupted strain. Furthermore, the final three amino acids of the peptide, which are critical for peroxisomal sorting, were not required for cross-linking to cyclophilin. We conclude that either cyclophilin is playing a nonessential facilitating role in peroxisomal targeting or that the interaction of the targeting peptide to cyclophilin is mimicking an interaction with an unidentified substrate or effector of cyclophilin.

scholarly journals An internal region of the peroxisomal membrane protein PMP47 is essential for sorting to peroxisomes

1994 ◽  
Vol 124 (6) ◽  
pp. 915-925 ◽  
Author(s):  
MT McCammon ◽  
JA McNew ◽  
PJ Willy ◽  
JM Goodman
Keyword(s):  
Amino Acids ◽  
Sequence Similarity ◽  
Candida Boidinii ◽  
Carrier Proteins ◽  
Peroxisomal Membrane ◽  
Lower Efficiency ◽  
Mitochondrial Carrier ◽  
Peroxisomal Targeting ◽  
Membrane Spanning ◽  
Mitochondrial Carrier Proteins

Targeting sequences on peroxisomal membrane proteins have not yet been identified. We have attempted to find such a sequence within PMP47, a protein of the methylotrophic yeast, Candida boidinii. This protein of 423 amino acids shows sequence similarity with proteins in the family of mitochondrial carrier proteins. As such, it is predicted to have six membrane-spanning domains. Protease susceptibility experiments are consistent with a six-membrane-spanning model for PMP47, although the topology for the peroxisomal protein is inverted compared with the mitochondrial carrier proteins. PMP47 contains two potential peroxisomal targeting sequences (PTS1), an internal SKL (residues 320-322) and a carboxy terminal AKE (residues 421-423). Using a heterologous in vivo sorting system, we show that efficient sorting occurs in the absence of both sequences. Analysis of PMP47-dihydrofolate reductase (DHFR) fusion proteins revealed that amino acids 1-199 of PMP47, which contain the first three putative membrane spans, do not contain the necessary targeting information, whereas a fusion with amino acids 1-267, which contains five spans, is fully competent for sorting to peroxisomes. Similarly, a DHFR fusion construct containing residues 268-423 did not target to peroxisomes while residues 203-420 appeared to sort to that organelle, albeit at lower efficiency than the 1-267 construct. However, DHFR constructs containing only amino acids 185-267 or 203-267 of PMP47 were not found to be associated with peroxisomes. We conclude that amino acids 199-267 are necessary for peroxisomal targeting, although additional sequences may be required for efficient sorting to, or retention by, the organelles.

scholarly journals Collagen cross-linking. Isolation of cross-linked peptides from collagen of chicken bone

1973 ◽  
Vol 135 (3) ◽  
pp. 393-403 ◽  
Author(s):  
D. R. Eyre ◽  
M. J. Glimcher
Keyword(s):  
Amino Acid ◽  
Bone Collagen ◽  
Collagen Molecule ◽  
Cross Linking ◽  
Bacterial Collagenase ◽  
Chicken Bone ◽  
Cross Links ◽  
The Cross ◽  
Significant Pathway ◽  
Carboxy Terminal

Cross-linked peptides were isolated from chicken bone collagen that had been digested with CNBr or with bacterial collagenase. Analyses of 3H radioactivity in disc electrophoretic profiles of the CNBr peptides from bone collagens that had been treated with NaB3H indicated that a major site of intermolecular cross-linking in chicken bone collagen is located between the carboxy-terminal region of an α1 chain and a small CNBr peptide, probably situated near the amino-terminus of an α1 or α2 chain in an adjacent collagen molecule. A small amount of this cross-linked CNBr peptide was isolated from a CNBr digest of chicken bone collagen by column chromatography. Amino acid analysis showed that the CNBr peptide, α1CB6B, the carboxy-terminal peptide of the α1 chain, was the major CNBr peptide in the preparation, and the reduced cross-linking components were identified as hydroxylysinohydroxynorleucine (HylOHNle), with a smaller amount of hydroxylysinonorleucine (HylNle). However, the composition and the low recovery of the cross-linking amino acids suggested that the preparation was a mixture of CNBr peptides α1CB6B and α1CB6B cross-linked to a small CNBr peptide whose identity could not be determined. A small cross-linked peptide was isolated from chicken bone collagen that had been reduced with NaB3H4 and digested with bacterial collagenase. This peptide was the major cross-linked peptide in the digest and contained a stoicheiometric amount of the reduced cross-linking compounds. A peptide which had the same amino acid composition, but contained the cross-linking compounds in their reducible forms, was isolated from a collagenase digest of chicken bone collagen that had not been treated with NaBH4. The absence of the reduced cross-links from this peptide indicates that, at least for the cross-linking site from which the peptide derives, natural reduction is not a significant pathway for biosynthesis of stable cross-links. However, most of the reducible cross-linking component in the peptide appeared to stabilize in the bone collagen by rearrangement from aldimine to ketoamine form.

scholarly journals Chemical compositions of elastins isolated from aortas and pulmonary tissues of humans of different ages

1972 ◽  
Vol 127 (1) ◽  
pp. 261-269 ◽  
Author(s):  
R. John ◽  
J. Thomas
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Cross Linking ◽  
Chemical Compositions ◽  
Amino Acid Compositions ◽  
Different Ages ◽  
Older Subjects ◽  
The Cross ◽  
Carbohydrate Contents

1. Elastins were isolated from the visceral pleuras and parenchymas of lungs of humans of different ages. 2. The elastin content of pleuras increased whereas that of parenchymas remained constant with increasing age. 3. The amino acid compositions and carbohydrate contents of elastins isolated from both pulmonary tissues changed in the same way with increasing age of the subjects. These changes were similar to those observed in elastins isolated from the aorta. 4. Similar glycoproteins were isolated from pleuras and aortas, and were more difficult to extract from the elastins of older subjects. Contamination with these glycoproteins was responsible for the changes in composition of elastin, as the age of the tissue from which it was extracted increased. 5. The amount of the cross-linking amino acids desmosine and isodesmosine was lower in elastins isolated from both aorta and pulmonary tissues of senile subjects than those from younger subjects.

scholarly journals Desmosine and isodesmosine contents and elastase activity in normal and cirrhotic rat liver

1983 ◽  
Vol 214 (3) ◽  
pp. 1023-1025 ◽  
Author(s):  
V Velebný ◽  
E Kasafírek ◽  
J Kanta
Keyword(s):  
Amino Acids ◽  
Carbon Tetrachloride ◽  
Rat Liver ◽  
Congo Red ◽  
Normal Liver ◽  
Cirrhotic Liver ◽  
Cross Linking ◽  
Elastase Activity ◽  
The Cross

The contents of desmosine and isodesmosine, the cross-linking amino acids of elastin, were increased 4-fold in rat liver with carbon tetrachloride-induced cirrhosis, which suggests that insoluble elastin accumulates in cirrhosis. Elastase activity in the cirrhotic liver, as determined with 3-carboxypropionyl-L-alanyl-L-alanyl-L-alanine p-nitroanilide, was 17% less than in the normal liver; no change was found when Congo Red-elastin was used as a substrate.

A photoenhanced oxidation of amino acids and the cross-linking of lysozyme mediated by tetrazolium salts

2021 ◽  
Vol 23 (6) ◽  
pp. 3761-3770
Author(s):  
Jianfeng Zhao ◽  
Ruixue Zhu ◽  
Xiting Zhang ◽  
Bowu Zhang ◽  
Yancheng Liu ◽  
...  
Keyword(s):  
Amino Acids ◽  
Uv Light ◽  
Cross Linking ◽  
Tetrazolium Salts ◽  
The Cross

Mechanisms of UV light-enabled strong oxidizing capacity of tetrazolium salts and their oxidization towards proteins were first elucidated.

Separation of the cross-linking amino acids of elastin on thin-layer plates

1984 ◽  
Vol 305 ◽  
pp. 461-464 ◽  
Author(s):  
Stephen Keller ◽  
Anjan K. Ghosh ◽  
Ajit K. Ghosh ◽  
Gerard M. Turino ◽  
Ines Mandl
Keyword(s):  
Amino Acids ◽  
Thin Layer ◽  
Cross Linking ◽  
The Cross
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