Protein chemical characterization of Mucor pusillus aspartic proteinase Amino acid sequence homology with the other aspartic proteinases, disulfide bond arrangement and site of carbohydrate attachment

AbstractThe amino acid sequence of Mucor pusillus aspartic protenaise was determined by analysis of fragments obtained from cleavage of the enzyme by CNBr and limited tryptic digestion. The protenaise is a single polypeptide chain protein containing 361 amino acid residues, cross-linked by two disulfide bonds. A sugar moiety composed of two GlcNAc residues and four neutral sugar residues is asparagine-linked to the chain. The sequence of M. pusillus proteinase is highly homologous with the M. miehei protonaise (83% identity). The homology with other aspartic proteinases is low (22–24%) and indicates that the Mucor proteinases diverged at an early evolutionary phase. The most conservative regions of the molecule are those involved in catalysis and forming the binding cleft and the core region of the molecule

De Novo Design of α-Amylase Inhibitor: A Small Linear Mimetic of Macromolecular Proteinaceous Ligands

SummaryWe report a low molecular weight inhibitor of α-amylases based on a linear peptidic scaffold designed de novo through the use of combinatorial chemistry. The inhibitory motif denoted PAMI (peptide amylase inhibitor) was selected by using L-peptide libraries and was fine-tuned by the introduction of unnatural modifications. PAMI specifically inhibits glycoside hydrolases of family 13. Its interaction with porcine pancreatic α-amylase was characterized by inhibition kinetics, fluorescence competition assays with natural α-amylase inhibitors, and isothermal titration calorimetry. We demonstrate that the critical amino acid residues in PAMI are shared with those in the macromolecular proteinaceous inhibitors that, however, bind to α-amylases through a spatially scattered set of intermolecular contacts. Thus, natural molecular evolution as well as combinatorial evolution selected the same α-amylase binding determinants for completely different spatial frameworks

Type 1 M protein of Streptococcus pyogenes N-terminal sequence and peptic fragments

AbstractLimited proteolysis of the surface of type 1 Streptococcus pyogenes by pepsin gives rise to fragment Pep M1 of Mr 20270 as the main product which covers the N-terminal part of the M protein. The amino acid sequence was determined of the N-terminal region of the M protein representing the most exposed part of the molecule on the surface fibrils of streptococcal cells, which seems to be very important for the differentiation of the individual serological types. The sequence differs from the homologous N-terminal sequences of types 5, 6 and 24, and shows a homology with sequences repeating in the chain of type 24. Fragment Pep M1 binds to fibrinogen; the absence of its 30 N-terminal amino acid residues, however, abolishes this interaction which is believed to play a role in the virulence of S. pyogenes