Chemical modification of peptides by hydrazine

High pressure (‘performance’) liquid chromatography on reverse-phase supports has been used to characterize the products arising from the hydrazine treatment of peptides. In addition to converting arginine residues into ornithine, the reaction was found to cleave predominately Gly-Xaa, Xaa-Gly, Asn-Xaa and Xaa-Ser peptide bonds. Peptide-bond cleavage and deguanidation was studied as a function of time of exposure to hydrazine, hydrazine concentration and temperature. The convenience of this method of chromatography for the rapid low-cost separation and isolation of peptides, as well as their reaction products, is illustrated at the level of material required for solid-phase microsequencing.

Download Full-text

Scandium(iii) triflate-promoted serine/threonine-selective peptide bond cleavage

Chemical Communications ◽

10.1039/c6cc10300f ◽

2017 ◽

Vol 53 (23) ◽

pp. 3311-3314 ◽

Cited By ~ 9

Author(s):

Jizhi Ni ◽

Youhei Sohma ◽

Motomu Kanai

Keyword(s):

Bond Cleavage ◽

Peptide Bond ◽

High Conversion ◽

Selective Hydrolysis ◽

Conversion Yield ◽

Peptide Bonds ◽

Peptide Bond Cleavage ◽

High Conversion Yield ◽

Site Selective ◽

Hydrolysis Of

The site-selective hydrolysis of peptide bonds at Ser and Thr positions was promoted by scandium(iii) triflate with a high conversion yield.

Download Full-text

Serine protease dynamics revealed by NMR analysis of the thrombin-thrombomodulin complex

Scientific Reports ◽

10.1038/s41598-021-88432-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Riley B. Peacock ◽

Taylor McGrann ◽

Marco Tonelli ◽

Elizabeth A. Komives

Keyword(s):

Active Site ◽

Serine Proteases ◽

Protein C ◽

Catalytic Mechanism ◽

Bond Cleavage ◽

Peptide Bond ◽

Peptide Bond Cleavage ◽

Exchange Mass Spectrometry ◽

Catalytically Active ◽

Hydrogen Deuterium

AbstractSerine proteases catalyze a multi-step covalent catalytic mechanism of peptide bond cleavage. It has long been assumed that serine proteases including thrombin carry-out catalysis without significant conformational rearrangement of their stable two-β-barrel structure. We present nuclear magnetic resonance (NMR) and hydrogen deuterium exchange mass spectrometry (HDX-MS) experiments on the thrombin-thrombomodulin (TM) complex. Thrombin promotes procoagulative fibrinogen cleavage when fibrinogen engages both the anion binding exosite 1 (ABE1) and the active site. It is thought that TM promotes cleavage of protein C by engaging ABE1 in a similar manner as fibrinogen. Thus, the thrombin-TM complex may represent the catalytically active, ABE1-engaged thrombin. Compared to apo- and active site inhibited-thrombin, we show that thrombin-TM has reduced μs-ms dynamics in the substrate binding (S1) pocket consistent with its known acceleration of protein C binding. Thrombin-TM has increased μs-ms dynamics in a β-strand connecting the TM binding site to the catalytic aspartate. Finally, thrombin-TM had doublet peaks indicative of dynamics that are slow on the NMR timescale in residues along the interface between the two β-barrels. Such dynamics may be responsible for facilitating the N-terminal product release and water molecule entry that are required for hydrolysis of the acyl-enzyme intermediate.

Download Full-text

Activation of Bovine Factor V by Thrombin and A Protease From Russell's Viper Venom (RVV)

10.1055/s-0039-1687642 ◽

1979 ◽

Author(s):

M.J. Lindhout ◽

C. M. Jackson

Keyword(s):

Bond Cleavage ◽

Peptide Bond ◽

Ion Exchange Chromatography ◽

Exchange Chromatography ◽

Factor V ◽

Peptide Bond Cleavage ◽

Prothrombinase Complex ◽

Factor Va ◽

Activation Products ◽

Ca Oxalate

In order to understand the function of activated factor V in the prothrombinase complex, we isolated the activation products obtained by action of thrombin and RVV-V on factor V and studied their functional properties. Factor V isolated from plasma by means of ion-exchange chromatography, a Ca-oxalate adsorption step and gelfiltration was homogenous in SDS-gelelectrophoresis (apparent MW 360,000, with and without reduction). Increase in factor V activity upon action by RVV-V is correlated with a single peptide bond cleavage, resulting in a 270,000 dalton and a 80,000 dalton component. Additional proteolysis of factor Va(RVV/V)’ by thrombin results in a further cleavage of the high MW component into peptides with MW's of 72,000, 94,000 and about 150,000 without a furth~r increase in factor V activity. Whereas none of the isolated peptides reveal factor Va activity, activity would be generated by a recombination in the presence of Ca2+ of the 94,000 MW or 270,000 MW component with the 80,000 component. Action of thrombin alone on factor V results in peptides of MW 72,000, 80,000, 94,000 and a peptide very rich in carbohydrate with an apparent MW of 150,000.

Download Full-text

The Protein Core of the Largest Proteoglycan Monomer of Articular Cartilage. Gel Electrophoretic Pattern and Tryptophanyl Peptide Bond Cleavage

Connective Tissue Research ◽

10.3109/03008208709005622 ◽

1987 ◽

Vol 16 (4) ◽

pp. 377-384 ◽

Cited By ~ 1

Author(s):

Victor Stanescu ◽

Françoise Chaminade ◽

Marie-Paule Muriel

Keyword(s):

Articular Cartilage ◽

Bond Cleavage ◽

Electrophoretic Pattern ◽

Peptide Bond ◽

Protein Core ◽

Peptide Bond Cleavage

Download Full-text

The Nature of Non-Specific Peptide Bond Cleavage During the Isothiocyanate Degradation of Proteins

Methods in Protein Sequence Analysis ◽

10.1007/978-1-4612-5832-2_6 ◽

1982 ◽

pp. 101-110 ◽

Cited By ~ 4

Author(s):

Wolf F. Brandt ◽

Agnes Henschen ◽

Claus Von Holt

Keyword(s):

Bond Cleavage ◽

Peptide Bond ◽

Peptide Bond Cleavage ◽

Specific Peptide

Download Full-text

Nitrogen-15 nuclear magnetic resonance spectroscopy. The state of histidine in the catalytic triad of .alpha.-lytic protease. Implications for the charge-relay mechanism of peptide-bond cleavage by serine proteases

Journal of the American Chemical Society ◽

10.1021/ja00494a001 ◽

1978 ◽

Vol 100 (26) ◽

pp. 8041-8047 ◽

Cited By ~ 196

Author(s):

William W. Bachovchin ◽

John D. Roberts

Keyword(s):

Nuclear Magnetic Resonance ◽

Magnetic Resonance ◽

Magnetic Resonance Spectroscopy ◽

Nuclear Magnetic Resonance Spectroscopy ◽

Serine Proteases ◽

Bond Cleavage ◽

Peptide Bond ◽

Catalytic Triad ◽

Resonance Spectroscopy ◽

Peptide Bond Cleavage

Download Full-text

The complete amino acid sequence of chicken skeletal-muscle enolase

Biochemical Journal ◽

10.1042/bj2360115 ◽

1986 ◽

Vol 236 (1) ◽

pp. 115-126 ◽

Cited By ~ 17

Author(s):

G A Russell ◽

B Dunbar ◽

L A Fothergill-Gilmore

Keyword(s):

Skeletal Muscle ◽

Amino Acid ◽

Amino Acid Sequence ◽

Peptide Bond ◽

Peptide Bonds ◽

Complete Amino Acid Sequence ◽

Arginine Residues ◽

Cnbr Cleavage ◽

Yeast Enolase ◽

Chicken Skeletal Muscle

The complete amino acid sequence of chicken skeletal-muscle enolase, comprising 433 residues, was determined. The sequence was deduced by automated sequencing of hydroxylamine-cleavage, CNBr-cleavage, o-iodosobenzoic acid-cleavage, clostripain-digest and staphylococcal-proteinase-digest fragments. The presence of several acid-labile peptide bonds and the tenacious aggregation of most CNBr-cleavage fragments meant that a commonly used sequencing strategy involving initial CNBr cleavage was unproductive. Cleavage at the single Asn-Gly peptide bond with hydroxylamine proved to be particularly useful. Comparison of the sequence of chicken enolase with the two yeast enolase isoenzyme sequences shows that the enzyme is strongly conserved, with 60% of the residues identical. The histidine and arginine residues implicated as being important for the activity of yeast enolase are conserved in the chicken enzyme. Secondary-structure predictions are analysed in an accompanying paper [Sawyer, Fothergill-Gilmore & Russell (1986) Biochem. J. 236, 127-130].

Download Full-text