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Journal Issue - Volume 2 Issue 2 (February 1993)

Abstract A ternary electron transfer protein complex has been crystallized and a preliminary structure investigation has been carried out. The complex is composed of a quinoprotein, methylamine dehydrogenase (MADH), a blue copper protein, amicyanin, and a c‐type cytochrome (c551i). All three proteins were isolated from Paracoccus denitrificans. The crystals of the complex are orthorhombic, space group C2221 with cell dimensions a = 148.81 Å, b...

Abstract The retinoblastoma gene product (Rb), a cellular growth suppressor, complexes with viral and cellular proteins that contain a specific binding domain incorporating three invariant residues: Leu‐X‐Cys‐X‐Glu, where X denotes a nonconserved residue. Hydrophobic and electrostatic properties are strongly conserved in this segment even though the nonconserved amino acids vary considerably from one Rb‐binding protein to another....

Abstract The three disulfide Ecballium elaterium trypsin inhibitor II (EETI II) reduction with dithiothreitol (DTT) and re‐oxidation of the fully reduced derivative have been examined. A common stable intermediate has been observed for both processes. Isolation and sequencing of carboxymethylated material showed that the intermediate lacks the [2–19] bridge. The NMR study showed a very strong structural conservation as compared to...

Abstract Crystallographic work on antigen‐antibody complexes has revealed that extensive surface areas of proteins may interact with antibodies. On the other hand, most experimental approaches to locate and define antigenic determinants of protein antigens rely on the linear sequence of the polypeptide chain. Hence the question arises whether mapping of antibody binding sites by analysis of the reactivity of anti‐protein antibodies...

Abstract A mass spectrometry method has been developed for selective detection of glycopeptides at the low (≤25) picomole level during chromatography of glycoprotein digests and for differentiation of O‐linked from N‐linked oligosaccharides. The technique involves observation of diagnostic sugar oxonium‐ion fragments, particularly the HexNAc+ fragment at m/z 204, from collisionally excited glycopeptides. Collision‐induced fragmentation can be accomplished...

Abstract Sau3 A and Hind III restriction fragments of Clostridium cylindrosporum genomic DNA were used to isolate clones containing 80% of the N10‐H4folate synthetase gene in a 5′ fragment and the remaining 20% of the gene in the 3′ fragment. These fragments were joined at a common SnaB I restriction site and expressed in Escherichia coli at a level equivalent to what is normally found in C. cylindrosporum. Sequence comparisons show a large degree of...

  • Modulation of antibody affinity by a non‐contact residue

  • Joel F. Schildbach, Richard I. Near, Robert E. Bruccoleri, Edgar Haber, Philip D. Jeffrey, Jiri Novotny, Steven Sheriff, Michael N. Margolies
  • Published in Wiley Interscience on Dec 31, 2008
  • DOI: 10.1002/pro.5560020209 (p 206-214)

Abstract Antibody LB4, produced by a spontaneous variant of the murine anti‐digoxin monoclonal antibody 26–10, has an affinity for digoxin two orders of magnitude lower than that of the parent antibody due to replacement of serine with phenylalanine at position 52 of the heavy chain variable region (Schildbach, J.F., Panka, D.J., Parks, D.R., et al., 1991, J. Biol. Chem. 266, 4640–4647). To examine the basis for the decreased affinity, a panel...

Abstract We analyzed sequences surrounding known tyrosine sulfation sites to determine the characteristics that distinguish these sites from those that do not undergo sulfation. Tests evaluated the number and position of acidic, basic, hydrophobic, and small amino acids, as well as disulfide and N‐glycosylation (sugar) sites. We determined that composition‐based tests that select close to 100% of known tyrosine sulfation sites...

Abstract The largest and the second‐largest subunit of the multisubunit eukaryotic RNA polymerases are involved in interaction with the DNA template and the nascent RNA chain. Using Southwestern DNA‐binding techniques and nitrocellulose filter binding assays of bacterially expressed fusion proteins, we have identified a region of the largest, 215‐kDa, subunit of Drosophila RNA polymerase II that has the potential to bind nucleic acids...

Abstract The nucleocapsid protein (NC) is the major genomic RNA binding protein that plays integral roles in the structure and replication of all animal retroviruses. In this report, select biochemical properties of recombinant Mason‐Pfizer monkey virus (MPMV) and HIV‐1 NCs are compared. Evidence is presented that two types of saturated Zn2 NC‐polynucleotide complexes can be formed under conditions of low [NaCl] that differ in apparent...

Abstract The folding pathway for a 150‐amino acid recombinant form of the dimeric cytokine human macrophage colony‐stimulating factor (M‐CSF) has been studied. All 14 cysteine residues in the biologically active homodimer are involved in disulfide linkages. The structural characteristics of folding intermediates blocked with iodoacetamide reveal a rapid formation of a small amount of a non‐native dimeric intermediate species...

Abstract GNA, the mannose‐specific lectin from Galanthus nivalis was confirmed to bind α‐2‐macroglobulin (A2M) but another protein was copurified with A2M from total human plasma. A total of 23 other lectins with diverse specificities were tested for reaction with human A2M and with three other members of the A2M family. NPA, a mannose‐specific lectin isolated from Narcissus pseudonarcissus bulbs, and RSA, the Rhizoctonia solani agglutinin,...

Abstract The family of aspartic proteinases includes several human enzymes that may play roles in both physiological and pathophysiological processes. The human lysosomal aspartic proteinase cathepsin D is thought to function in the normal degradation of intracellular and endocytosed proteins but has also emerged as a prognostic indicator of breast tumor invasiveness. Presented here are results from a continuing effort to elucidate...

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