Site-directed mutagenesis of glyceraldehyde-3-phosphate dehydrogenase reveals the role of residue Ser148

A mutant of Bacillus stearothermophilus D-glyceraldehyde-3-phosphatedehydrogenase, Ser¹⁴⁸ – Ala, was produced byoligonucleotide-directedmutagenesis. The study of the catalytic properties of this mutanthas shown that this mutation significantly affects the Michaelisconstant of inorganic phosphate and to a lesser extent thatof 1,3-diphosphoglycerate and D-glyceraldehyde-3-phosphate.This result is consistent with model-building studies whichshow that, for the phosphorylation step of catalysis, inorganicphosphate must bind to the anion recognition site designatedP_i with the C(3) phosphate of the acyl-enzyme intermediate inthe alternative anion site P_s. Studies of the enantiomeric specificityusing D- and L-glyceraldehyde as substrates show that the hydroxylgroup of Ser¹⁴⁸, combined with the presence of the C(3) phosphateof the substrate, enhances stereospecificity as well as catalysis.However, the stereospecific effect cannot be a consequence ofthe direct interaction of Ser¹⁴⁸ with the C(2)-hydroxyl of thesubstrate. The changed K_m for glyceraldehyde-3-phosphate suggeststhat the initial step of hemithioacetal formation may take placewith its C(3) phosphate bound in the P_i site. This supportsthe molecular mechanism proposed by Moody (1984). Therefore,catalysis could be enhanced through interactions of the serinehydroxyl group not only with inorganic phosphate but also withthe C(3) phosphate of glyceraldehyde-3-phosphate.     

Use of site-directed mutagenesis to probe the role of Cys149 in the formation of charge-transfer transition in glyceraldehyde-3-phosphate dehydrogenase

Oligonucleotide-directed mutagenesis was employed to producemutants of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH)of Escherichia coli and Bacillus stearothermophilus. Three differentmutant proteins—His¹⁷⁶ — Asn, Cys¹⁴⁹ — Ser,Cys¹⁴⁹ — Gly—were isolated from one or both of theenzymes. The study of the properties of these mutants has shownthat Cys¹⁴⁹ is clearly responsible for the information of acharge-transfer transition, named the Racker band, observedduring the NAD⁺ binding to apoGAPDH. This result excludes asimilarity between the Racker band and the charge-transfer transitionobserved following the alkylation of GAPDH by 3-chloroacetylpyridine-adenine dinucleotide.     

Distributed enterprise information networking: A case example

A major health care provider is migrating its existing centralized computing system to a distributed computing infrastructure that will support advanced clinical, business, and workgroup applications and connectivity. Deploying a modern distributed architecture integrated with a traditional, heterogeneous mainframe-oriented enterprise data processing and information retrieval environment is an enormous undertaking. This process requires major changes for the enterprise, especially in the areas of the network, control and operations, systems deployment, integration and cost management tools, and personnel. Our case example is Kaiser Permanente Northern California Region (KPNCR). Information Technology Services (ITS), a division of KPNCR, provides network and computer support and operation, architecture and standards, and information technology. This division, in partnership with business planning and operational units, is responsible for creating the distributed architecture. The transition, estimated to take three to five years, is underway. ITS plans to deploy the proposed architecture at three medical facilities. This pilot will provide the technology foundation for a concurrent pilot of a planned enterprise-wide clinical information system. Results from the pilot will influence plans to deploy the distributed architecture enterprise-wide     

Self-appraisal, life satisfaction, and retrospective life choices across one and three decades.

This research investigated the relationship of a self-appraisal of having lived up to one's intellectual abilities at midlife (average age of 49 years) with life satisfaction and retrospective life choices 1 and 3 decades later among 383 participants in the Terman Study of the Gifted. Study 1 showed that participants who reported living up to their intellectual abilities were higher in satisfaction with occupational success, satisfaction with family life, and joy in living 11 years later. Study 2 showed that participants who reported living up to their abilities were higher in overall life satisfaction and were less likely to report that they would make different life choices in work or family life 3 decades later. In an integrative structural equation model, the relation between the midlife self-appraisal of having lived up to intellectual abilities and overall satisfaction at age 80 was mediated by life satisfaction discrepancy at age 61. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The x-ray crystal structure of phosphomannose isomerase from Candida albicans at 1.7 angstrom resolution

Phosphomannose isomerase (PMI) catalyses the reversible isomerization of fructose-6-phosphate (F6P) and mannose-6-phosphate (M6P). Absence of PMI activity in yeasts causes cell lysis and thus the enzyme is a potential target for inhibition and may be a route to antifungal drugs. The 1.7 A crystal structure of PMI from Candida albicans shows that the enzyme has three distinct domains. The active site lies in the central domain, contains a single essential zinc atom, and forms a deep, open cavity of suitable dimensions to contain M6P or F6P The central domain is flanked by a helical domain on one side and a jelly-roll like domain on the other.