Investigation of subtilisin digest of pepsin chain between half-cystines II and III

1981 ◽  
Vol 46 (3) ◽  
pp. 640-654
Author(s):  
Vladimír Kostka
Keyword(s):  
Amino Acid ◽  
Paper Chromatography ◽  
Sequential Analysis ◽  
Cysteine Residue ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Hydrolysis Of ◽  
Dowex 1 ◽  
Final Purification

Aminoethylated hog pepsin was subjected to tryptic digestion and the longest fragment, arising from cleavage at S-(β-aminoethyl)-cysteine residue No II and III was isolated from the digest. This fragment was subjected to additional cleavage with subtilisin and the digest resolved into crude fractions by chromatography on Dowex 1. The isolation and final purification of the peptides was carried out by paper electrophoreses and paper chromatography. By these methods 55 peptides were obtained which were subjected to sequential analysis by stepwise degradation. The amino acid sequences of these peptides and their position in the pepsin chain are given. These sequences provided overlaps for peptides obtained by hydrolysis of this part of the pepsin chain by other enzymes.

scholarly journals Haemoglobins of the Shark, Heterodontus Portusjacksoni II. Amino Acid Sequence of the a-Chain

1976 ◽  
Vol 29 (2) ◽  
pp. 73 ◽  
Author(s):  
AR Nash ◽  
WK Fisher ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Paper Chromatography ◽  
Cyanogen Bromide ◽  
Gel Filtration ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Amino Terminal ◽  
Core Peptide ◽  
A Chain

The amino acid sequence of the a-chain of the principal haemoglobin from the shark, H. portusjacksoni has been determined. The chain has 148 residues and is acetylated at the amino terminal. The soluble peptides obtained by tryptic and chymotryptic digestion of the protein or its cyanogen bromide fragments were isolated by gel filtration, paper ionophoresis and paper chromatography. The amino acid sequences were determined by the dansyl-Edman procedure. The insoluble 'core' peptide from the tryptic digestion contained 34 residues and required cleavage by several proteases before the sequence was established. Compared with human a-chain there are 88 amino acid differences including the additional seven residues which appear on the amino terminal of the shark chain. There is also one deletion and one insertion. The chain contains no tryptophan but has four cysteinyl residues which is the highest number of such residues recorded for a vertebrate globin.

scholarly journals 1,2-α-d-mannosidase from Penicillium citrinum: molecular and enzymic properties of two isoenzymes

1993 ◽  
Vol 290 (2) ◽  
pp. 349-354 ◽  
Author(s):  
T Yoshida ◽  
T Inoue ◽  
E Ichishima
Keyword(s):  
Amino Acid ◽  
Gel Permeation Chromatography ◽  
Amino Acid Sequences ◽  
Penicillium Citrinum ◽  
Tryptic Digestion ◽  
Terminal Amino Acid ◽  
Sds Page ◽  
Terminal Amino ◽  
Hydrolysis Of ◽  
Similar Kinetic

Two isoforms of acidic 1,2-alpha-D-mannosidases have been isolated from culture filtrate of Penicillium citrinum. The pI values of the two forms, designated 1,2-alpha-mannosidase Ia and Ib, were 4.6 and 4.7 respectively. Isoenzymes Ia and Ib exhibited the same molecular mass which was determined to be 53 kDa by SDS/PAGE and 54 kDa by gel-permeation chromatography. Enzymes Ia and Ib hydrolysed yeast mannan and 1,2-alpha-linked mannooligosaccharides, but did not hydrolyse p-nitrophenyl alpha-D-mannoside. The optimal pH for the hydrolysis of Man(alpha 1-->2)Man was 5.0 for both isoenzymes. Similar kinetic parameters were determined for the two forms. Activation energy was a little lower for Ia than Ib. There was little difference between the enzymes with regard to their performance at acidic or alkaline pH. The N-terminal amino acid sequences of the two enzymes were identical. Analysis of C-terminal peptides, which were prepared by tryptic digestion and anhydrotrypsin-agarose chromatography, showed that Ia and Ib had the same amino acid sequences in the C-terminal region. Tryptic digestion revealed a slight difference between the isoenzymes in the pattern of cleaved peptides on SDS/PAGE.

scholarly journals The Fine Structure of the Sensory Epithelium of the Vomeronasal Organ in Suckling Rats

1971 ◽  
Vol 24 (3) ◽  
pp. 765 ◽  
Author(s):  
Jean E Kratzing
Keyword(s):  
Amino Acid ◽  
Gel Filtration ◽  
Vomeronasal Organ ◽  
Sensory Epithelium ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Tryptic Peptides ◽  
Special Procedure ◽  
A Chain

The amino acid sequence of the a-chain of haemoglobin from M. giganteus has been determined. The soluble peptides formed by tryptic digestion were isolated by gel filtration, ion-exchange chromatography, paper ionophoresis, and chromatography. The amino acid sequences were determined by the "dansyl"Edman procedure. Incomplete hydrolysis of one bond resulted in a large insolublecore peptide containing 40 amino acid residues. The sequence of this peptide was deduced from the sequences of smaller peptides resulting from further digestion with thermolysin and papain. Maleylation of the a-globin before tryptic digestion gave three large fragments which assisted in assigning tryptic peptides to specific areas of the molecule. A special procedure involving maleylation of a chymotryptic digest of globin was used to isolate peptides containing arginine which provided overlap sequences of tryptic peptides

ISOLATION AND STRUCTURE OF URIDINE NUCLEOTIDE-PEPTIDES FROM AEROBACTER CLOACAE NRC 492

1963 ◽  
Vol 41 (1) ◽  
pp. 1065-1072 ◽  
Author(s):  
R. A. Anwar ◽  
C. Roy ◽  
R. W. Watson
Keyword(s):  
Lactic Acid ◽  
Amino Acid ◽  
Lithium Chloride ◽  
Paper Chromatography ◽  
High Voltage ◽  
Chloride Solutions ◽  
Uridine Nucleotide ◽  
Ethanol Extracts ◽  
Dowex 1 ◽  
High Voltage Electrophoresis

Four closely related uridine nucleotide-peptides isolated from ethanol extracts of penicillin-treated Aerobacter cloacae NRC 492 have been partially characterized. Fractionation of dialyzed extracts on Dowex-1-Cl columns with aqueous lithium chloride solutions, precipitation of the nucleotide-peptides with methanol–acetone, and further separation by paper chromatography and high voltage electrophoresis yielded (a) UDP-GNAc-lact.L-ala.D-glu.meso-dap.D-ala,* (b) UDP-GNAc-lact.L-ala.D-glu.meso-dap, (c) UDP-GNAc-lact.L-ala.D-glu., and (d) UDP-GNAc-lactic acid. The amino acid isomers were identified by micro-enzymic and chromatographic methods.

scholarly journals Purification of two distinct types of phosphoinositide-specific phospholipase C from rat liver. Enzymological and structural studies

1988 ◽  
Vol 256 (2) ◽  
pp. 453-459 ◽  
Author(s):  
O Nakanishi ◽  
Y Homma ◽  
H Kawasaki ◽  
Y Emori ◽  
K Suzuki ◽  
...  
Keyword(s):  
Amino Acid ◽  
Phospholipase C ◽  
Rat Liver ◽  
Peptide Mapping ◽  
Polyacrylamide Gel Electrophoresis ◽  
Liver Homogenate ◽  
Amino Acid Sequences ◽  
Dependent Manner ◽  
Phosphatidylinositol 4 ◽  
Hydrolysis Of

Two kinds of phosphoinositide-specific phospholipase C (PLC) were purified from rat liver by acid precipitation and several steps of column chromatography. About 50% of the activity could be precipitated when the pH of the liver homogenate was lowered to pH 4.7. The redissolved precipitate yielded two peaks, PLC I and PLC II, in an Affi-gel Blue column, and each was further purified to homogeneity by three sequential h.p.l.c. steps, which were different for the two enzymes. The purified PLC I and PLC II had estimated Mr values of 140,000 and 71,000 respectively on SDS/polyacrylamide-gel electrophoresis. Both enzymes hydrolysed phosphatidylinositol (PI), phosphatidylinositol 4-phosphate (PIP) and phosphatidylinositol 4,5-bisphosphate (PIP2) in a Ca2+- and pH-dependent manner. PLC I was most active at 10 microM- and 0.1 mM-Ca2+ for hydrolysis of PI and PIP2 respectively, whereas PLC II showed the highest activity at 5 mM- and 10 microM-Ca2+ for that of PI and PIP2 respectively. The optimal pH of the two enzymes also differed with substrates or Ca2+ concentration, in the range pH 5.0-6.0. Hydrolysis of phosphoinositides by these enzymes was completely inhibited by Hg2+ and was affected by other bivalent cations. From data obtained by peptide mapping and partial amino acid sequencing, it was clarified that PLC I and PLC II had distinct structures. Moreover, partial amino acid sequences of three proteolytic fragments of PLC I completely coincided with those of PLC-148 [Stahl, Ferenz, Kelleher, Kriz & Knopf (1988) Nature (London) 332, 269-272].

scholarly journals Phenotypic and Enzymatic Comparative Analysis of the Novel KPC Variant KPC-5 and Its Evolutionary Variants, KPC-2 and KPC-4

2008 ◽  
Vol 53 (2) ◽  
pp. 557-562 ◽  
Author(s):  
Daniel J. Wolter ◽  
Philip M. Kurpiel ◽  
Neil Woodford ◽  
Marie-France I. Palepou ◽  
Richard V. Goering ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Single Amino Acid ◽  
Upstream Region ◽  
The Novel ◽  
Carbapenem Resistant ◽  
Flanking Regions ◽  
Hydrolysis Of ◽  
Additional Amino Acid

ABSTRACT A novel Klebsiella pneumoniae carbapenemase (KPC) variant, designated bla KPC-5, was discovered in a carbapenem-resistant Pseudomonas aeruginosa clinical isolate from Puerto Rico. Characterization of the upstream region of bla KPC-5 showed significant differences from the flanking regions of other bla KPC variants. Comparison of amino acid sequences with those of other KPC enzymes revealed that KPC-5 was an intermediate between KPC-2 and KPC-4, differing from KPC-2 by a single amino acid substitution (Pro103→Arg), while KPC-4 contained Pro103→Arg plus an additional amino acid change (Val239→Gly). Transformation studies with an Escherichia coli recipient strain showed differences in the properties of the KPC variants. KPC-4 and KPC-5 both had pIs of 7.65, in contrast with the pI of 6.7 for KPC-2. KPC-2 transformants were less susceptible to the carbapenems than KPC-4 and KPC-5 transformants. These data correlated with higher rates of imipenem hydrolysis for KPC-2 than for KPC-4 and KPC-5. However, KPC-4 and KPC-5 transformants had higher ceftazidime MICs, and the enzymes from these transformants had slightly better hydrolysis of this drug than KPC-2. KPC-4 and KPC-5 were more sensitive than KPC-2 to inhibition by clavulanic acid in both susceptibility testing and hydrolysis assays. Thus, KPC enzymes may be evolving through stepwise mutations to alter their spectra of activity.

scholarly journals Studies on Monotreme Proteins. V. Amino Acid Sequence of the a-Chain of Haemoglobin From the Platypus, Ornithorhynchus anatinus

1974 ◽  
Vol 27 (6) ◽  
pp. 591 ◽  
Author(s):  
RG Whittaker ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Complete Amino Acid Sequence ◽  
Ornithorhynchus Anatinus ◽  
A Chain

Blood from the platypus contained three haemoglobins which were separated by chromatography on DEAE-Sephadex. The major component, Hb-I, was converted to globin and fractionated into the oc-and p-chains by chromatography on eM-cellulose in 8M urea-thiol buffers, and the complete amino acid sequence of the 141 residues of the oc-chain were determined. Peptides derived from the oc-chain by tryptic digestion were isolated by paper ionophoresis and chromatography. The amino acid sequences were determined by the dansyl-Edman procedure or by further digestion with other enzymes.

scholarly journals Studies on Monotreme Proteins II. Amino Acid Sequence of the a-Chain in Haemoglobin From the Echidna, Tachyglossus Aculeatus Aculeatus

1973 ◽  
Vol 26 (4) ◽  
pp. 877 ◽  
Author(s):  
RG Whittaker ◽  
WK Fisher ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Paper Chromatography ◽  
Gel Filtration ◽  
Tryptic Digestion ◽  
A Chain ◽  
Tachyglossus Aculeatus

The amino acid sequence of the 141 residues of the IX-chain of the major haemoglobin (Hb-IB) from the echidna has been determined. The soluble peptides formed by tryptic digestion were isolated by gel filtration, paper ionophoresis, and paper chromatography.

scholarly journals Studies on Marsupial Protein. II. Amino Acid Sequence of the -Chain of Haemoglobin from the Grey Kangaroo, Macropus Giganteus

1969 ◽  
Vol 22 (6) ◽  
pp. 1437 ◽  
Author(s):  
GM Air ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Sequence ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Grey Kangaroo

The amino acid sequence of the jS-chain of haemoglobin from M. giganteus has been determined. The soluble peptides formed by tryptic digestion were isolated by gel filtration, ion-exchange chromatography, and paper ionophoresis, and amino acid sequences determined by the "dansyl"-Edman procedure. Special procedures were necessary for three peptides which were insoluble.

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