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Journal Issue - Volume 1 Issue 4 (April 1992)

  • Progress and innovation

  • Hans Neurath
  • Published in Wiley Interscience on Dec 31, 2008
  • DOI: 10.1002/pro.5560010401 (p 425-425)

Abstract Thrombin is a multifunctional serine proteinase that plays a key role in coagulation while exhibiting several other key cellular bioregulatory functions. The X‐ray crystal structure of human α‐thrombin was determined in its complex with the specific thrombin inhibitor d‐Phe‐Pro‐Arg chloromethylketone (PPACK) using Patterson search methods and a search model derived from trypsinlike proteinases of known spatial structure (Bode, W.,...

Abstract The allosteric transition of glycogen phosphorylase promoted by protein phosphorylation is accompanied by the association of a pair of functional dimers to form a tetramer. The conformational changes within the dimer that lead to the creation of a protein recognition surface have been analyzed from a comparison of the crystal structures of T‐state dimeric phosphorylase b and R‐state tetrameric phosphorylase a. Regions of...

Abstract We report the design, chemical synthesis, and structural and functional characterization of a novel reagent for protein sequence analysis by the Edman degradation, yielding amino acid derivatives rapidly detectable at high sensitivity by ion‐evaporation mass spectrometry. We demonstrate that the reagent 3‐[4′(ethylene‐N, N, N‐trimethylamino)phenyl]‐2‐isothiocyanate is chemically stable and shows coupling and cyclization/cleavage yields...

Abstract Calmodulin from Saccharomyces cerevisiae was expressed in Escherichia coli and purified. The purified protein was structurally characterized using limited proteolysis followed by ESI mass spectrometry to identify the fragments. In the presence of Ca2+, yeast calmodulin is sequentially cleaved at arginine 126, then lysine 115, and finally at lysine 77. The rapid cleavage at Arg‐126 suggests that the fourth Ca2+‐binding loop does not...

Abstract Arg‐127 stabilizes the oxyanion of the tetrahedral intermediate formed during Zn2+ carboxypeptidase A‐catalyzed hydrolysis. Mutant carboxypeptidases lacking Arg‐127 exhibit substantially reduced rates of hydrolysis with the change manifest almost entirely in kcat (kcat/Km is decreased by 104 for R127A). Therefore, Arg‐127 stabilizes the enzyme‐transition state complex but not the ground state enzyme‐substrate complex (Phillips, M.A.,...

Abstract Citrate synthase (CS), which has been denatured in either guanidine hydrochloride (GdnHCl) or urea can be assisted in its renaturation in a variety of ways. The addition of each of the assistants – bovine serum albumin (BSA), oxaloacetate (OAA), and glycerol – promotes renaturation. In combination, the effect of these substances is additive with respect to the yield of folded CS. The report of Buchner et al. (Buchner, J.,...

Abstract The ability of high pressure to dissociate several peripheral protein‐membrane complexes was investigated. Three vitamin K‐dependent proteins (factor X, protein Z, and prothrombin) dissociated from small unilamellar vesicles (SUVs, 30 nm diameter) composed of 25% phosphatidylserine (PS) and 75% phosphatidylcholine (PC) at comparable pressures (midpoints of 0.3–0.6 kbar). The pressure‐induced dissociation curves for the...

Abstract Flavocytochrome b2 (or l‐lactate dehydrogenase) from baker's yeast is thought to operate by the initial formation of a carbanion, as do the evolutionarily related α‐hydroxy acid‐oxidizing FMN‐dependent oxidases. Previous work has shown that, in the active site of the unligated reduced flavocytochrome b2, the group that has captured the substrate α‐proton has a high pKapp, calculated to lie around 15 through the use of Eigen's equation....

Abstract EntF is the enzyme responsible for serine activation during the biosynthesis of enterobactin (a cyclic trimer of N‐dihydroxybenzoyl serine) in Escherichia coli. EntF has been overexpressed and purified to >90% homogeneity. The enzyme has been shown to complement the entF− MK1 strain in the synthesis of 2,3‐dihydroxybenzoylserine derivatives and exhibits l‐serine‐dependent ATP[32P] pyrophosphate exchange activity with a Km for serine of 260 mM and a...