Site‐directed mutagenesis has been used to produce mutant forms of yeast phosphoglycerate kinase in which the conserved active‐site residue, Arg21, has been replaced by a methionine or a lysine. Kinetic results obtained using these mutant enzymes show that their K m for both 3‐phospho‐D‐glycerate and ATP are significantly different from those recorded for the wild‐type enzyme. The V max for the lysine mutant is reduced by a factor of two from that of the wild‐type enzyme whereas the V max for the methionine mutant is reduced more than sevenfold. A very clean electron‐density‐difference map shows little, if any, evidence of a structural change associated with the C‐terminal domain, although resonances in the NMR spectra associated with the ATP‐binding site (C‐terminal domain) are also affected by the mutation as one might expect from the kinetic results. The NMR data show that binding at both the 3‐phospho‐D‐glycerate and the non‐productive ATP‐binding site (associated with the N‐terminal domain) are affected in the mutant in a way which is different to that associated with the wild‐type enzyme. These results, taken together with the X‐ray and kinetic data, indicate that the non‐productive ATP‐binding site and the activating anion‐binding site are both associated with the basic patch region of yeast phosphoglycerate kinase.

Phosphoglycerate kinase

Site-directed mutagenesis

Wild type

Residue (chemistry)

10.1111/j.1432-1033.1992.tb17016.x

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Citations (8)

THE GLYCERATE MUTASE PHOSPHORYL TRANSFER SYSTEM

Biochemical Society Transactions (1981)

H.C. Watson Linda A. Fothergill

Conference Abstract| April 01 1981 THE GLYCERATE MUTASE PHOSPHORYL TRANSFER SYSTEM H. C. Watson; H. C. Watson *Department of Biochemistry, University of Bristol, BS8 1TD, England, U.K. Search for other works by this author on: This Site PubMed Google Scholar L. A. Fothergill L. A. Fothergill +Department of Biochemistry, University of Aberdeen, AB9 1AS, Scotland, U.K. Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1981) 9 (2): 20P. https://doi.org/10.1042/bst009020pa Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation H. C. Watson, L. A. Fothergill; THE GLYCERATE MUTASE PHOSPHORYL TRANSFER SYSTEM. Biochem Soc Trans 1 April 1981; 9 (2): 20P. doi: https://doi.org/10.1042/bst009020pa Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search This content is only available as a PDF. © 1981 Biochemical Society1981 Article PDF first page preview Close Modal You do not currently have access to this content.

Watson

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10.1042/bst009020pa

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Citations (0)

Molecular Symmetry of Glyceraldehyde-3-phosphate Dehydrogenase

WORLD SCIENTIFIC eBooks (2014)

Michael G. Rossmann Geoffrey C. Ford H.C. Watson Leonard Banaszak

Glyceraldehyde 3-phosphate dehydrogenase

Glyceraldehyde

10.1142/9789814513357_0012

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Citations (0)

Low Resolution Structure of Yeast Phosphoglycerate Kinase

Nature New Biology (1972)

P.L. Wendell T. N. Bryant H.C. Watson

Phosphoglycerate kinase

Phosphoglycerate mutase

10.1038/newbio240134a0

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Conformational Changes and Inhibitor Binding at the Active Site of Elastase

Cold Spring Harbor Symposia on Quantitative Biology (1972)

David M. Shotton Nicholas J. White H.C. Watson

Elastase is one of a class of serine proteinases (Hartley, 1960) which also includes trypsin, chymotrypsin-A and chymotrypsin-B, enzymes which all exhibit a common catalytic mechanism for the hydrolysis of peptide bonds in dietary protein, but which differ markedly in their specificity towards the amino acid residues forming those bonds. Elastase is synthesized in the acinar tissue of mammalian pancreas as an inactive zymogen, proelastase, which on entering the duodenum is activated by the tryptic cleavage of a single peptide bond, releasing a small activation peptide (Gertler and Birk, 1970) to yield the active enzyme, a single polypeptide chain of 240 amino acid residues containing four disulfide bridges (Shotton and Hartley, 1970).

Zymogen

Peptide bond

Pancreatic elastase

Active center

10.1101/sqb.1972.036.01.015

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Citations (39)

Structure of Deoxymyoglobin : A Crystallographic Study

Nature (1966)

C. L. Nobbs H.C. Watson J Kendrew

10.1038/209339a0

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Citations (138)

Atomic coordinates for tosyl-elastase

Biochemical and Biophysical Research Communications (1973)

Lindsay Sawyer David M. Shotton H.C. Watson

Tosyl

10.1016/0006-291x(73)90183-6

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Citations (16)

Structure and activity of phosphoglycerate mutase

Philosophical transactions of the Royal Society of London. Series B, Biological sciences (1981)

SUSAN I. WINN H.C. Watson Richard N. Harkins Linda A. Fothergill

The structure of yeast phosphoglycerate mutase determined by X-ray crystallographic and amino acid sequence studies has been interpreted in terms of the chemical, kinetic and mechanistic observations made on this enzyme. There are two histidine residues at the active site, with imidazole groups almost parallel to each other and approximately 0.4 nm apart, positioned close to the 2 and 3 positions of the substrate. The simplest interpretation of the available information suggests that a ping-pong type mechanism operates in which at least one of these histidine residues participates in the phosphoryl transfer reaction. The flexible C-terminal region also plays an important role in the enzymic reaction.

Phosphoglycerate mutase

Phosphoglycerate kinase

Imidazole

Mutase

10.1098/rstb.1981.0066

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Citations (107)