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A purified glycoprotein with blood-group-A specificity, was digested with insoluble pronase to give a serologically active product enriched in peptide-bonded serine and threonine residues; this was degraded with 0.25 M sulphuric in glacial acetic acid for 24 h. The resultant N-acetylgalactosaminyl-peptide were hydrolysed with 0.2 M hydrocholoric acid for 24 h at 110 °C. fractionation of the products on Sephadex G-10, followed by preparative separation on amino-acid analyzer and paper electrophoresis at pH 1.9 yielded the following homogeneous oligopeptides; one tripeptide with the N-terminal sequence Thr-Thr-Ser, three tetrapeptides with the sequences, Thr-Ser-Thr-Ser, Thr-Pro-Thr-Ser and Pro-Thr-Thr-Ser, one pentapeptide with the sequence Thr-Pro-Thr-Ser, one hexapeptide with the sequence Pro-Thr-Thr-Thr-Pro-Ser and one heptapeptide with the sequence Ala-Pro-Thr-Thr-Ser-Gly-Ser. Galactosamine was present in several of these fragments and the hexapeptide and heptapeptide both contained sufficient of this sugar for all the serine and threonine residues to be glycosylated. The following five tetrapeptides sequences were deduced from mixtures of two peptides which differed by the amino-acid substituent at only one position in the chain; Pro-Thr-Ala-ser, Thr-Thr Pro-Ser, thr-Thr-Ala-Ser, Ser-Pro-Thr-Ser, and Ser-Ala-Thr-Ser. A tripeptide mixture which differed only in the nature of the third amino acid yielded the following three tripeptide sequences: Pro-Thr-Ser, Pro-thr-Ala. These peptides do not indicate a clearly defined amino-acid sequence surrounding the glycosylated hydroxy amino acids but establish that in the human blood-group-specific glycoproteins there is close packing of the threonine in the region of the peptide moiety carrying the carbohydrate chains. [ABSTRACT FROM AUTHOR]

The globin of sheep heart myoglobin was first cleaved by cyanogen bromide and the resulting fragments were fractionated by gel filtration and preparative paper electrochromatography; each fragment was submitted to endopeptidase digestion and the complete amino acid sequence has been determined. The intact globin and the median cyanogen bromide peptide were analysed by the protein sequanator (Edman). This analysis confirmed the sequence of the first 46 N- terminal residues of the intact globin and established the sequence of the first 39 N-terminal residues of the median cyanogen bromide peptide. Between sheep and beef myoglobin six differences over 153 residues can be noticed; among the six amino acid replacements, two correspond to the exchange of two bases and four to the exchange of one base in the coding triplets. [ABSTRACT FROM AUTHOR]

The αA2 chain of bovine α-crystallin was fragmented by means of cyanogen bromide treatment and by tryptic, chymotryptic and thermolytic digestions. Twenty tryptic peptides were obtained from the S-aminoethylated αA2 chain, accounting together for the complete sequence. The direct Edman degradation and the dansyl-Edman technique were used to determine the sequences of the tryptic peptides. The order of the tryptic peptides was deduced from overlapping peptides obtained by cyanogen bromide treatment, tryptic digestion of the maleylated chain and chymotryptic and thermolytic digestion of the S-aminoethylated chain. The sequence of the αA2 chain comprises 173 residues and corresponds to a molecular weight of 19832. [ABSTRACT FROM AUTHOR]

The protein part of phospholipase A2 from the venom of the common European honey-bee consists of a single chain of 128 amino acid residues [l], eight of which are half-cystine. The absence of free sulphydryl groups indicates that the enzyme contains four disulphide bridges. Digestion of the native enzyme with pepsin resulted in formation of a fragment containing all four disulphide bridges. Further digestion of this fragment with trypsin allowed the isolation of two singly cross-linked peptides, one containing half-cystine residues 37 and 107, and the other containing residues 59 and 99. A fragment containing the remaining two disulphide bridges was digested with dilute sulphuric acid and a peptide was isolated from the digest in which half-cystine residues 9 and 30 were linked. The remaining disulphide bridge is between residues 31 and 89. [ABSTRACT FROM AUTHOR]

The complete amino acid sequence of neurotoxin I of the North African scorpion Androctonus australis Hector collected in Algeria (area of Chellala) is described. S-Methylation of cysteine residues formed by reduction of disulphide bonds was found useful for their identification in sequence determination using the phenylisothiocyanate method. From the N-terminal sequence (38 residues) established in the protein sequenator, and from the sequence of chymotryptic peptides the complete sequence of the protein has been deduced. The molecule consists of a single polypeptide chain cross-linked by four disulphide bridges. An isoneurotoxin (I') has been found in the venom of A. australis collected in Tunisia (area of Tozeur). It differs from neurotoxin I in the replacement, of the valine residue in position 17 by an isoleucine residue. [ABSTRACT FROM AUTHOR]

One Bence Jones protein and two Bence Jones fragments (fragmentary chains) of K type, and one Bence Jones protein and two fragments of L type have been analyzed. These proteins have been aminoethylated and digested with proteolytic enzymes. The resulting peptides were separated by different procedures. The peptides obtained were analyzed and could be located in most. cases in the amino acid sequence of the corresponding protein by homology with published sequences. These tentative sequences were particularly satisfactory in the case of the three K type proteins. These belonged to subtype ICE and showed considerable homologies with other KI proteins. It was more difficult to position peptides of the three L type proteins on the basis of homology with published sequences. Several peptides of one protein (BJ 98) which were completely sequenced by Edman degradation combined with proteolytic digestion could not be positioned. Amino acid analysis of peptides obtained from Bence Jones fragments confirmed that these correspond to the N-terminal variable region of Bence Jones proteins for K type fragments, whereas they extend for three residues over the C-terminal constant region for some L type fragments. In any case, no peptide from the constant region of light chains has been isolated from tryptic digests of Bence Jones fragments. [ABSTRACT FROM AUTHOR]

The λ-chains of normal pig γG-immunoglobulin were digested with pepsin in 1 M formic acid. Cystine-containing peptides were isolated by gel filtration on Sephadex G-25 and by chromatography on Dowex 50 and Dowex 1. The following structures were found:[this equation cannot be presented in ASCII text] In addition to these peptides obtained in a high yield similar peptides were found but their complete structures were not determined in view of their low yield. The structures found in the λ-chains display a high degree of similarity with corresponding structures known to exist in human Bence­Jones proteins of the L-type. [ABSTRACT FROM AUTHOR]

The complete amino acid sequence of neurotoxin II of the North African scorpion Androctonus auatratis Hector has been determined. From the N-terminal sequence (43 residues) obtained by automatic phenylisothiocyanate degradation in the protein sequenator, and from the sequence of tryptic peptides obtained by digestion of the reduced and methylated toxin of which amino groups had been previously masked by reaction with ethylthioltrifluoroacetate the complete amino acid sequence of the protein has been inferred. It is composed of a single polypeptide chain of 64 amino acid residues, including 8 half-cystines, ended by an histidinamide residue at the C-terminus. [ABSTRACT FROM AUTHOR]

Histone F3 has been fragmented into 10 non-overlapping fragments. With the aid of the automatic Edman degradation procedure for proteins using 3-(N,N-dimethylamino)propyne buffer the first 48 amino-acid residues in the uncleared protein have been positioned. The remainder of the amino-acid sequence of this protein was elucidated by subjecting the aligned peptides to the automatic degradation. The amino acids obtained during the degradation were quantitated and identified by gas chromatography and amino-acid analysis. The primary structure of this histone is compared to the corresponding fraction from calf which had been independently determined by another laboratory. [ABSTRACT FROM AUTHOR]

3-Phosphoglyeerate kinase has been purified for the first time from a glueoneogenetic tissue. A seven-step procedure for the preparation of the bovine-liver enzyme, which results in a specific activity of 130-150 units/mg, is described. The overall purification was 250-fold and 17% of the original activity was recovered. About 50 mg pure protein can be obtained from 1 kg liver tissue. Optimum conditions for crystallization and stabilization of the enzyme have been elaborated. The existence of at least three different forms of phosphoglycerate kinase in liver extracts has been demonstrated by isoelectric focussing experiments. The main activity peak was purified to homogeneity. Evidence was obtained for a probable tissue specificity of the isoelectric points of the known phosphoglycerate-kinase enzymes (bovine liver pH 6.5, pig liver 6.3; rabbit muscle 7.0; yeast 7.2; mice Ehrlich ascites cells 6.8). Bovine-liver phosphoglycerate kinase was shown to be homogeneous by sedimenting as a regular sharp peak in the analytical ultracentrifuge and by the appearance of a single protein band on both cellulose acetate and polyacrylamide disc gel electrophoresis. The liver and yeast-type enzymes were all distinguishable by their migration distances at pH 4.9, 7.0 and 8.6. Homogeneity of the bovine-liver enzyme was further confirmed by rechromatography on Biogel P-150 and by the determination of the unique NH2- and COOH-terminal amino-acid sequences NH2-His-Lys-Tyr and Thr-Glu-COOH, respectively. The results obtained demonstrate that liver phosphoglycerate kinase, purified by the method described, is a homogeneous protein, distinct from the known muscle and yeast enzymes. [ABSTRACT FROM AUTHOR]

The determination of the amino acid sequence of the C-peptide of human proinsulin is presented. Two constituent fragments of C-peptide, Chy 1+ 2 and Chy 3 (residues 1-24 amd 25-31, respectively) were liberated by mild chymotryptic digestion and isolated gel filtration and ion exchange chromatography. From Chy 1 + 2 was obtained seven smaller peptides by further digestion with chymotrypsin. The sequence analysis of these fragments allowed an overall structure to be assigned. Papain digestion of the intact C-peptide provided 12 peptides, the compositions of which provided unequivocal confirmation for the proposed sequence. [ABSTRACT FROM AUTHOR]

Acidic proteins can be removed from calf thymus deoxyribonucleoprotein, previously washed with 0.14 M NaCl, using 0.35 M NaCl. After the removal of these acidic proteins, the yield of acidic nuclear proteins as extracted by other methods dropped to less than one third. Calf thymus deoxyribonucleoprotein and reconstituted deoxyribonucleohistone adsorb acidic proteins from the cytoplasm and nuclear sap in 0.14 M NaCl. These proteins which can be released in 0.35 M NaCl are similar in amino acid analysis to the acidic nuclear proteins prepared by other method. [ABSTRACT FROM AUTHOR]

Chloroform-soluble proteins have been isolated free from phosphatides from rat liver mitochondria, microsomes and cytosol by chromatography on Sephadex LH-20 columns. The amino acid composition of the chloroform-soluble and insoluble proteins has been estimated. The chloroform-soluble proteins from mitochondria and cytosol have a higher content of hydrophobic and a lower content of acidic amino acids compared to the corresponding chloroform-insoluble proteins. No marked differences were found between chloroform-soluble and insoluble proteins from microsomes. In contrast to the acidic chloroform-insoluble proteins, all chloroform-soluble proteins are basic. From the amino acid composition the amount of chloroform-soluble proteins has been estimated. Mitochondrial membranes contain 1.6%, mitochondrial matrix proteins less than 0.025%, microsomes 3.0% and the cytosol 0.15% from total proteins. The molecular weight distribution of chloroform-soluble proteins synthesized in mitochondria has been measured by gelelectrophoresis. The molecular weights range from about 7000 to 30000. The turnover of chloroform-soluble proteins from mitochondria and cytosol is higher than from the corresponding chloroform-insoluble proteins. A part of mitochondrial chloroform-soluble proteins (about 30%) is synthesized within the mitochondria. [ABSTRACT FROM AUTHOR]

Reduced, S-aminoethylated guinea-pig α-lactalbumin was cleaved with eyanogen bromide at its single methionyl residue. The products were separated gel by filtration and characterized by amino acid analysis and N-terminal group determination, as a 90-residue peptide from the N-terminal of the protein (N-terminal lysine) and a 33-residue peptide from the C-terminus of the protein (N-terminal aminnethyl cysteine). The amino acid sequences of both these peptides were determined by examination of the peptides obtained by digestion of the purified fragments with trypsin, ehymotrypain or thermotysin. The complete amino acid sequence of guinea-pig α-lactalbumin thus obtained is discussed in relation to the known sequences of other members of the Iysozyme-α-lactalbumin group. [ABSTRACT FROM AUTHOR]