On the three-dimensional structure and catalytic mechanism of triose phosphate isomerase

1981 ◽  
Vol 293 (1063) ◽  
pp. 159-171 ◽  
Keyword(s):  
Catalytic Mechanism ◽  
Polypeptide Chain ◽  
Three Dimensional ◽  
Triose Phosphate Isomerase ◽  
Dimensional Structure ◽  
Dihydroxyacetone Phosphate ◽  
Triose Phosphate ◽  
Independent Determination ◽  
Chicken Muscle

Triose phosphate isomerase is a dimeric enzyme of molecular mass 56000 which catalyses the interconversion of dihydroxyacetone phosphate (DHAP) and D-glyceraldehyde-3-phosphate. The crystal structure of the enzyme from chicken muscle has been determined at a resolution of 2.5 A, and an independent determination of the structure of the yeast enzyme has just been completed at 3 A resolution. The conformation of the polypeptide chain is essentially identical in the two structures, and consists of an inner cylinder of eight strands of parallel |3-pleated sheet, with mostly helical segments connecting each strand. The active site is a pocket containing glutamic acid 165, which is believed to act as a base in the reaction. Crystallographic studies of the binding of DHAP to both the chicken and the yeast enzymes reveal a common mode of binding and suggest a mechanism for catalysis involving polarization of the substrate carbonyl group.

scholarly journals Enzyme-substrate and enzyme-inhibitor complexes of triose phosphate isomerase studied by 31P nuclear magnetic resonance

1979 ◽  
Vol 179 (3) ◽  
pp. 607-621 ◽  
Author(s):  
I D Campbell ◽  
R B Jones ◽  
P A Kiener ◽  
S G Waley
Keyword(s):  
Enzyme Inhibitor ◽  
Ph Dependence ◽  
Triose Phosphate Isomerase ◽  
Dihydroxyacetone Phosphate ◽  
Ligand Complex ◽  
Triose Phosphate ◽  
Kinetic Information ◽  
Transition State Analogue ◽  
Chicken Muscle ◽  
Proton Uptake

The complex formed between the enzyme triose phosphate isomerase (EC 5.3.1.1.), from rabbit and chicken muscle, and its substrate dihydroxyacetone phosphate was studied by 31P n.m.r. Two other enzyme-ligant complexes examined were those formed by glycerol 3-phosphate (a substrate analogue) and by 2-phosphoglycollate (potential transition-state analogue). Separate resonances were observed in the 31P n.m.r. spectrum for free and bound 2-phosphoglycollate, and this sets an upper limit to the rate constant for dissociation of the enzyme-inhibitor complex; the linewidth of the resonance assigned to the bound inhibitor provided further kinetic information. The position of this resonance did not vary with pH but remained close to that of the fully ionized form of the free 2-phosphoglycollate. It is the fully ionized form of this ligand that binds to the enzyme. The proton uptake that accompanies binding shows protonation of a group on the enzyme. On the basis of chemical and crystallographic information [Hartman (1971) Biochemistry 10, 146–154; Miller & Waley (1971) Biochem. J. 123, 163–170; De la Mare, Coulson, Knowles, Priddle & Offord 1972) Biochem. J. 129, 321–331; Phillips, Rivers, Sternberg, Thornton & Wilson (1977) Biochem. Soc. Trans. 5, 642–647] this group is believed to be glutamate-165. On the other hand, the position of the resonance of D-glycerol 3 phosphate (sn-glycerol 1-phosphate) in the enzyme-ligand complex changes with pH, and both monoanion and dianon of the ligand bind, although dianion binds better. The substrate, dihydroxyacetone phosphate, behaves essentially like glycerol 3-phosphate. The experiments with dihydroxy-acetone phosphate and triose phosphate isomerase have to be carried out at 1 degree C because at 37 degrees C there is conversion into methyl glyoxal and orthophosphate. The mechanismof the enzymic reaction and the reasons for rate-enhancement are considered, and aspects of the pH-dependence are discussed in an Appendix.

scholarly journals Specificity and kinetics of triose phosphate isomerase from chicken muscle

1972 ◽  
Vol 129 (2) ◽  
pp. 301-310 ◽  
Author(s):  
Sylvia J. Putman ◽  
A. F. W. Coulson ◽  
I. R. T. Farley ◽  
B. Riddleston ◽  
J. R. Knowles
Keyword(s):  
Triose Phosphate Isomerase ◽  
Proton Exchange ◽  
The Other ◽  
Chicken Breast ◽  
Dihydroxyacetone Phosphate ◽  
Exchange Reactions ◽  
Hydrogen Atoms ◽  
Triose Phosphate ◽  
Chicken Muscle ◽  
Kinetics Of

The isolation of crystalline triose phosphate isomerase from chicken breast muscle is described. The values of kcat. and Km for the reaction in each direction were determined from experiments over wide substrate-concentration ranges, and the reactions were shown to obey simple Michaelis–Menten kinetics. With d-glyceraldehyde 3-phosphate as substrate, kcat. is 2.56×105min-1and Km is 0.47mm; with dihydroxyacetone phosphate as substrate, kcat. is 2.59×104min-1and Km is 0.97mm. The enzyme-catalysed exchange of the methyl hydrogen atoms of the ‘virtual substrate’ monohydroxyacetone phosphate with solvent2H2O or3H2O was shown. This exchange is about 104-fold slower than the corresponding exchange of the C-3 hydrogen of dihydroxyacetone phosphate. The other deoxy substrate, 3-hydroxypropionaldehyde phosphate, was synthesized, but is too unstable in aqueous solution for analogous proton-exchange reactions to be studied.

An Analysis of the Three-Dimensional Structure of Chicken Triose Phosphate Isomerase

1977 ◽  
Vol 5 (3) ◽  
pp. 642-647 ◽  
Author(s):  
D. C. PHILLIPS ◽  
P. S. RIVERS ◽  
M. J. E. STERNBERG ◽  
J. M. THORNTON ◽  
I. A. WILSON
Keyword(s):  
Three Dimensional ◽  
Triose Phosphate Isomerase ◽  
Dimensional Structure ◽  
Three Dimensional Structure ◽  
Triose Phosphate

The three-dimensional structure of crystalline porcine pancreatic elastase

1970 ◽  
Vol 257 (813) ◽  
pp. 111-118 ◽  
Keyword(s):  
Amino Acid ◽  
Catalytic Mechanism ◽  
Polypeptide Chain ◽  
Molecular Model ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Side Chains ◽  
Peptide Bonds ◽  
Single Polypeptide Chain ◽  
Single Polypeptide

Elastase is a proteolytic enzyme obtained from pig pancreas, which shows a high degree of amino acid sequence homology with other serine proteinases, including bovine trypsin and chymotrypsin (Hartley, this volume, p. 77). It consists of a single polypeptide chain of 240 residues, which corresponds to the single polypeptide chain of trypsin, and the B and C chains of chymotrypsin. Elastase possesses a common catalytic mechanism with these enzymes but differs from them in its substrate specificity, cleaving peptide bonds on the carboxyl terminal side of amino acid residues lacking charged or aromatic side chains (Naughton & Sanger 1961). Several workers have suggested that homologous enzymes with common catalytic mechanisms have very similar tertiary structures. This prediction was supported by Blow and his co-workers, who found that the two disulphide bridges present in trypsin, but absent in chymotrypsin, could be built into the molecular model of a-chymotrypsin with little or no distortion of the polypeptide chain (Sigler, Blow, Matthews & Henderson 1968), and by Hartley (this volume, p. 77) who has shown that the trypsin and elastase side chains can be substituted for those present in a skeletal molecular model of a-chymotrypsin with no gross distortions of the polypeptide chain.

Conformation of Ribosomes from Escherichia Coli

1974 ◽  
Vol 32 ◽  
pp. 210-211
Author(s):  
M. Boublik ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Binding Sites ◽  
Ribosomal Proteins ◽  
Fluorescent Dyes ◽  
Protein Biosynthesis ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Dimensional Structure ◽  
Complete Understanding ◽  
And Function

The present knowledge of the three-dimensional structure of ribosomes is far too limited to enable a complete understanding of the various roles which ribosomes play in protein biosynthesis. The spatial arrangement of proteins and ribonuclec acids in ribosomes can be analysed in many ways. Determination of binding sites for individual proteins on ribonuclec acid and locations of the mutual positions of proteins on the ribosome using labeling with fluorescent dyes, cross-linking reagents, neutron-diffraction or antibodies against ribosomal proteins seem to be most successful approaches. Structure and function of ribosomes can be correlated be depleting the complete ribosomes of some proteins to the functionally inactive core and by subsequent partial reconstitution in order to regain active ribosomal particles.

The first crystal structure of the peptidase domain of the U32 peptidase family

2015 ◽  
Vol 71 (12) ◽  
pp. 2505-2512 ◽  
Author(s):  
Magdalena Schacherl ◽  
Angelika A. M. Montada ◽  
Elena Brunstein ◽  
Ulrich Baumann
Keyword(s):  
Crystal Structure ◽  
Catalytic Mechanism ◽  
Quaternary Structure ◽  
Catalytic Domain ◽  
Three Dimensional ◽  
Zinc Ion ◽  
Crystal Structure Analysis ◽  
Dimensional Structure ◽  
Sequence Motifs ◽  
Conserved Sequence

The U32 family is a collection of over 2500 annotated peptidases in the MEROPS database with unknown catalytic mechanism. They mainly occur in bacteria and archaea, but a few representatives have also been identified in eukarya. Many of the U32 members have been linked to pathogenicity, such as proteins fromHelicobacterandSalmonella. The first crystal structure analysis of a U32 catalytic domain fromMethanopyrus kandleri(genemk0906) reveals a modified (βα)8TIM-barrel fold with some unique features. The connecting segment between strands β7 and β8 is extended and helix α7 is located on top of the C-terminal end of the barrel body. The protein exhibits a dimeric quaternary structure in which a zinc ion is symmetrically bound by histidine and cysteine side chains from both monomers. These residues reside in conserved sequence motifs. No typical proteolytic motifs are discernible in the three-dimensional structure, and biochemical assays failed to demonstrate proteolytic activity. A tunnel in which an acetate ion is bound is located in the C-terminal part of the β-barrel. Two hydrophobic grooves lead to a tunnel at the C-terminal end of the barrel in which an acetate ion is bound. One of the grooves binds to aStrep-Tag II of another dimer in the crystal lattice. Thus, these grooves may be binding sites for hydrophobic peptides or other ligands.

scholarly journals Determination of the alpha-actinin-binding site on actin filaments by cryoelectron microscopy and image analysis.

1994 ◽  
Vol 126 (2) ◽  
pp. 433-443 ◽  
Author(s):  
A McGough ◽  
M Way ◽  
D DeRosier
Keyword(s):  
Image Analysis ◽  
Smooth Muscle ◽  
Binding Sites ◽  
Actin Filaments ◽  
Three Dimensional ◽  
Actin Binding ◽  
Dimensional Structure ◽  
Outer Face ◽  
Actin Binding Domain

The three-dimensional structure of actin filaments decorated with the actin-binding domain of chick smooth muscle alpha-actinin (alpha A1-2) has been determined to 21-A resolution. The shape and location of alpha A1-2 was determined by subtracting maps of F-actin from the reconstruction of decorated filaments. alpha A1-2 resembles a bell that measures approximately 38 A at its base and extends 42 A from its base to its tip. In decorated filaments, the base of alpha A1-2 is centered about the outer face of subdomain 2 of actin and contacts subdomain 1 of two neighboring monomers along the long-pitch (two-start) helical strands. Using the atomic model of F-actin (Lorenz, M., D. Popp, and K. C. Holmes. 1993. J. Mol. Biol. 234:826-836.), we have been able to test directly the likelihood that specific actin residues, which have been previously identified by others, interact with alpha A1-2. Our results indicate that residues 86-117 and 350-375 comprise distinct binding sites for alpha-actinin on adjacent actin monomers.

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