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K Biemann

Publications and source records attributed to K Biemann.

At least 55 records · Page 3Linked to original sources
PubMed1989

Glutaredoxin from rabbit bone marrow. Purification, characterization, and amino acid sequence determined by tandem mass spectrometry.

A glutaredoxin was purified from rabbit bone marrow, and its amino acid sequence was determined by high performance tandem mass spectrometry. The sequences of peptides generated by digestion with trypsin alone or in combination with thermolysin were determined from their collision-induced dissociation (CID) mass spectra. Alignment of these sequences and additional sequence information were obtained from the collision-induced dissociation mass spectra of peptides obtained from digestion of the intact protein with Staphylococcus aureus V8 protease and alpha-chymotrypsin. The resulting sequence of 106 amino acids is as follows: Ac-Ala-Gln-Glu-Phe-Val-Asn-Ser-Lys-Ile-Gln-Pro-Gly-Lys-Val-Val-Val-Phe- Ile-Lys-Pro-Thr-Cys-Pro-Tyr-Cys-Arg-Lys-Thr-Gln-Glu-Ile-Leu-Ser-Glu-Leu- Pro-Phe - Lys-Gln-Gly-Leu-Leu-Glu-Phe- Val-Asp-Ile-Thr-Ala-Thr-Ser-Asp-Met-Ser-Glu-Ile- Gln-Asp-Tyr-Leu-Gln-Gln-Leu-Thr-Gly-Ala-Arg- Thr-Val-Pro-Arg-Val-Phe-Leu-Gly-Lys-Asp-Cys-Ile- Gly-Gly-Cys-Ser-Asp-Leu-Ile-Ala-Met-Gln-Glu-Lys- Gly-Glu-Leu-Leu-Ala-Arg-Leu-Lys-Glu-Met-Gly- Ala-Leu-Arg-Gln. This glutaredoxin strongly resembles the corresponding calf and pig proteins (known as glutaredoxin and thioltransferase, respectively) with respect to its primary structure and enzymatic activity as a GSH:disulfide thioltransferase, an activity also found for the glutaredoxin from Escherichia coli. However, rabbit glutaredoxin was not active as a hydrogen donor for the reduction of ribonucleotides in the presence of the ribonucleotide reductases from rabbit bone marrow, Lactobacillus leichmannii, and Corynebacterium nephridii.

Amino Acid Sequence↗
PubMed1989

A revised sequence of calf thymus glutaredoxin.

The previously published structure of the glutaredoxin from calf thymus [Klintrot et al., (1984) Eur. J. Biochem. 144, 417-423] was reinvestigated by tandem mass spectrometry and found to have an N-terminal Ac-Ala-Gln-Ala-... sequence and an additional four amino acids inserted between positions 67 and 68.

Amino Acid Sequence↗
PubMed1989

The structure of tyrosine aminotransferase. Evidence for domains involved in catalysis and enzyme turnover.

The primary structure of tyrosine aminotransferase, as deduced from the nucleotide sequence of complementary DNA, was confirmed by fast atom bombardment mass spectrometry of tryptic peptides derived from the purified protein. Limited digestion of the native enzyme with trypsin released an acetylated, amino-terminal peptide; the new amino terminus in the modified enzyme was Val65. Endogenous proteases generated a chromatographically separable form of tyrosine aminotransferase that began at Lys35. Neither trypsin nor the other proteases altered the catalytic activity of tyrosine aminotransferase. Reduction of the holoenzyme with sodium borohydride yielded a major tryptic peptide containing phosphopyridoxamine bound to lysine 280, which probably functions in transamination. The carboxyl terminus of tyrosine aminotransferase contains features that typify proteins with short half-lives; it includes two negatively charged, hydrophilic segments that are enriched for glutamyl residues and are similar to a PEST region in ornithine decarboxylase (Rogers, S., Wells, R., and Rechsteiner, M. (1986) Science 234, 364-368). Tyrosine aminotransferase belongs to a superfamily of enzymes which includes aspartate aminotransferase and can be aligned so that many invariant, functional residues coincide. Like the isoenzymes of aspartate aminotransferase, tyrosine aminotransferase may contain two domains, with a central, catalytic core, and a small domain made up of both amino- and carboxyl-terminal components. We speculate that the exposed small domain may confer the unusually rapid degradative rate that characterizes this enzyme.

Amino Acid Sequence↗
PubMed1989

Computer program (SEQPEP) to aid in the interpretation of high-energy collision tandem mass spectra of peptides.

An algorithm (SEQPEP) that aids in the interpretation of high-energy (greater than 1 kV) collision-induced dissociation mass spectra of peptide ions generated by fast atom bombardment (FAB) ionization is described. The only required input is a list of product ion masses and relative abundances generated by the mass spectrometer data system, the mass of the precursor [M + H]+ ion, and the mass of any C-terminal modification, if present (e.g., amide). Possible N-terminal modifications and amino acid compositions are not required as input. In the output, sequences are ranked according to the fraction of total product ion current that can be accounted for as either sequence-specific or non-sequence-specific fragment ions. These are listed by ion type. One of the major advantages of this program over algorithms described earlier is the incorporation of ion types more recently discovered. Also, this program is much faster, requiring less than 5 min of central processing unit time for an input of as many as 100 product ions. The results obtained from 50 peptides, including some generated when sequencing a protein of previously unknown structure, are discussed.

Amino Acid Sequence↗
PubMed1988

Amino acid sequence of thioredoxin isolated from rabbit bone marrow determined by tandem mass spectrometry.

The amino acid sequence of the thioredoxin isolated from rabbit bone marrow was determined chiefly by high performance tandem mass spectrometry and fast atom bombardment mass spectrometry combined with manual Edman degradation. The sequences of peptides generated by digestion with trypsin alone or in combination with Staphylococcus aureus protease V8 or thermolysin were determined from their collision-induced dissociation mass spectra. Alignment of these sequences and additional sequence information were obtained from the collision-induced dissociation mass spectra of peptides obtained from digestion of the intact protein with S. aureus protease V8 and alpha-chymotrypsin. The resulting sequence of 104 residues is as follows: Val-Lys-Gln-Ile-Glu-Ser-Lys-Ser-Ala-Phe-Gln- Glu-Val-Leu-Asp-Ser-Ala-Gly-Asp-Lys-Leu-Val-Val- Val-Asp-Phe-Ser-Ala-Thr-Trp-Cys-Gly-Pro-Cys-Lys- Met-Ile-Lys-Pro-Phe-Phe-His-Ala-Leu-Ser-Glu-Lys- Phe-Asn-Asn-Val-Val-Phe-Ile-Glu-Val-Asp-Val-Asp- Asp-Cys-Lys-Asp-Ile-Ala-Ala-Glu-Cys-Glu-Val-Lys- Cys-Met-Pro-Thr-Phe-Gln-Phe-Phe-Lys-Lys- Gly-Gln-Lys-Val-Gly-Glu-Phe-Ser-Gly-Ala-Asn-Lys- Glu-Lys-Leu-Glu-Ala-Thr-Ile-Asn-Glu-Leu-Leu.

Amino Acid Sequence↗
PubMed1988

Characterization of the glycosylation sites in yeast external invertase. I. N-linked oligosaccharide content of the individual sequons.

External invertase is the product of the SUC2 gene of Saccharomyces cerevisiae. The deduced sequence of this enzyme (Taussig, R., and Carlson, M. (1983) Nucleic Acid Res. 11, 1943-1954) reveals it to contain 14 potential N-linked glycosylation sites, or sequons, although only 9-10 appear to be glycosylated (Trimble, R. B., and Maley, F. (1977) J. Biol. Chem. 252, 4409-4412). To determine the location of the glycosylated sequons, external invertase was deglycosylated with endo-beta-acetylglucosaminidase H and its component peptides analyzed by both fast atom bombardment mass spectrometry (FABMS) and classical peptide isolation procedures. By use of the former technique most of the glucosamine-containing sequons could be located and by the latter sufficient amounts of small glucosamine-containing peptides were isolated to enable their quantitation. From the combined FABMS and glucosamine analyses, it was established that eight of the sequons in a subunit of invertase are either completely or almost completely glycosylated, while five others are glycosylated to the extent of about 50% or less. In the case of two overlapping sequons (4 and 5), which include Asn92-Asn93-Thr-Ser, only the first Asn was glycosylated. Thus, all but one of the sequons of external invertase are glycosylated to some extent, giving an appearance of only 9-10 N-linked oligosaccharides/subunit. The sequence identity of both external and internal invertase was verified by FABMS and by peptide sequence analysis. In only one site was an amino acid found to differ from that deduced from the DNA sequence of the SUC2 gene. This occurred at position 390 where a proline was found in place of alanine, which could result from a single base change in the triplet specifying the latter amino acid.

Amino Acid Sequence↗
PubMed1988

Contributions of mass spectrometry to peptide and protein structure.

With the advent of recombinant deoxyribonucleic acid (DNA) technology which allows a wide range of manipulation of genes and their expression in cell lines other than the natural ones, many aspects of protein structure have become more important than ever. In addition to the determination of the amino acid sequence questions relating to homogeneity, the nature of post-translational modifications, the verification of the structure of a protein produced by a synthetically modified gene or the detection of a natural mutant are all questions that are more and more frequently asked and with the demand for more detail. Mass spectrometry has emerged as an important contributor to this field, particularly since the advent of fast atom bombardment (FAB) ionization, which makes it possible to ionize directly large polar molecules such as peptides and small proteins. As such, FAB mass spectrometry provides mainly molecular weight information which, in itself, often suffices to answer certain questions, particularly because the mass of many peptides can be determined directly from a single mass spectrum of a mixture. However, in order to obtain detailed structural information, such as the amino acid sequence, fragmentation has to be induced by collision processes and the product ions separated, preferably in the second mass spectrometer of a tandem system. This approach is particularly suited for the determination of the sequence of N-blocked peptides and the nature of the blocking group; the type and location of modified (i.e. phosphorylated, sulfated, glycosylated) amino acids; detection or verification of amino acid replacements; confirmation of the structure of synthetic peptides; and last but not least, the determination of the primary structure of proteins.

Amino Acid Sequence↗
PubMed1988

Thioredoxin from Rhodospirillum rubrum: primary structure and relation to thioredoxins from other photosynthetic bacteria.

Thioredoxin was isolated from a photosynthetic purple nonsulfur bacterium, Rhodospirillum rubrum, and its primary structure was determined by high-performance tandem mass spectrometry. The sequence identity of R. rubrum thioredoxin to Escherichia coli thioredoxin was intermediate to those of the Chlorobium thiosulfatophilum and Chromatium vinosum proteins. The results indicate that R. rubrum has an NADP-thioredoxin system similar to that of other photosynthetic purple bacteria.

Amino Acid Sequence↗
PubMed1987

Characterization by tandem mass spectrometry of structural modifications in proteins.

Tandem mass spectrometry can be used to solve a number of protein structural problems that are not amenable to conventional methods for amino acid sequencing. Typical problems that use this approach involve characterization of peptides with blocked amino termini or peptides that have been otherwise posttranslationally processed, such as, by phosphorylation or sulfation. The structure and homogeneity of synthetic peptides can also be evaluated. Since peptides can be selectively characterized in the presence of other peptides or contaminants, the need for extensive purification is reduced or eliminated.

Amino Acid Sequence↗
PubMed1987

Fast atom bombardment mass spectrometry and tandem mass spectrometry of biologically active peptidoglycan monomers from Neisseria gonorrhoeae.

Fast atom bombardment mass spectrometry (FABMS) and tandem mass spectrometry (MS/MS) were employed to define the structures of Neisseria gonorrhoeae peptidoglycan monomers that were of interest because of their abilities to mediate diverse biological reactions ranging from arthritogenicity to somogenicity. FABMS-determined molecular weights of individual components present in several different enzymatically derived classes of gonococcal monomers revealed that each of these classes was a complex mixture of up to 13 distinct peptidoglycan fragments. These ranged from the predominant disaccharide tetrapeptides possessing reducing or nonreducing 1,6-anhydro-N-acetylmuramic acid ends to relatively minor constituents containing glycine or asparagine in addition to traditional peptidoglycan amino acids, i.e. alanine, glutamic acid, and diaminopimelic acid. FABMS of high performance liquid chromatography-purified monomers yielded some sequence information; however, analysis even of unfractionated peptidoglycan mixtures using a JEOL HX110/HX110 tandem mass spectrometer operating at 10 kV provided unambiguous primary sequence data for the peptidoglycan monomers and defined the position of glycine in four compounds as well as the location of O-acetyl substituents (present on some compounds) on C-6 of the N-acetylmuramic acid residue.

Amino Acid Sequence↗
PubMed1987

Mass spectrometrically derived amino acid sequence of thioredoxin from Chlorobium, an evolutionarily prominent photosynthetic bacterium.

The amino acid sequence of the thioredoxin isolated from the photosynthetic green sulfur bacterium Chlorobium thiosulfatophilum was determined chiefly by fast atom bombardment mass spectrometry combined with Edman degradation and tandem mass spectrometry. For this purpose, the protein was digested with trypsin, alpha-chymotrypsin, thermolysin, and Staphylococcus aureus protease or combinations thereof. Chemical cleavage with cyanogen bromide was also used alone or in combination with trypsin. The resulting sequence of 108 amino acids is as follows: Ala-Gly- Lys-Tyr-Phe-Glu-Ala-Thr-Asp-Lys-Asn-Phe-Gln- Thr-Glu-Xle-Xle-Asp-Ser-Asp-Lys-(Ala-Val)-Xle- Val-Asp-Phe-Trp-Ala-Ser-Trp-Cys-Gly-(Pro-Cys)- Met-Met-Xle-Gly-Pro-Val-Xle-Glu-Gln-Xle-Ala-Asp- Asp-Tyr-Glu-Gly-Lys-Ala-Xle-Xle-Ala-Lys-Xle-Asn- Val-Asp-Glu-Asn-Pro-Asn-Xle-Ala-Gly-Gln-Tyr-Gly- Xle-Arg-Ser-Xle-Pro-Thr-Met-Xle-Xle-Xle-Ly s- (Gly-Gly-Lys)-Val-Val-Asp-Gln-Met-Val-Gly-Ala- Xle-Pro-Lys-Asn-Met-Xle-Ala-Lys-Lys-Xle-Asp-Glu-His-Il e-Gly (where Xle represents leucine or isoleucine; sequences in parentheses are based on homology considerations). It exhibits less than 53% homology with Escherichia coli thioredoxin.

Amino Acid Sequence↗
PubMed1987

The primary structure of thioredoxin from Chromatium vinosum determined by high-performance tandem mass spectrometry.

The primary structure of thioredoxin, a redox protein isolated from Chromatium vinosum, was determined by high-performance tandem mass spectrometry, which permitted sequencing of the 14 peptides (ranging in length from 2 to 18 amino acids) generated by digestion with trypsin and of several peptides produced by Staphylococcus aureus protease. The mass spectrometrically determined molecular weights of the peptides from the latter digest were used to properly align the tryptic peptides, which could also be accomplished on the basis of the considerable homology with Escherichia coli thioredoxin. Finally, the molecular weight of the Chromatium thioredoxin was determined by mass spectrometry and found to be 11,748.0, in good agreement with 11,750.2 calculated for the proposed sequence. Although it was difficult to establish by mass spectrometry, five leucines and three isoleucines could be identified, leaving only eight undifferentiated.

Amino Acid Sequence↗