Kinetic Features (kinetic + feature)

Distribution by Scientific Domains

Selected Abstracts

Kinetic and biochemical analyses on the reaction mechanism of a bacterial ATP-citrate lyase

FEBS JOURNAL, Issue 14 2002
Tadayoshi Kanao
The prokaryotic ATP-citrate lyase is considered to be a key enzyme of the carbon dioxide-fixing reductive tricarboxylic acid (RTCA) cycle. Kinetic examination of the ATP-citrate lyase from the green sulfur bacterium Chlorobium limicola (Cl -ACL), an ,4,4 heteromeric enzyme, revealed that the enzyme displayed typical Michaelis-Menten kinetics toward ATP with an apparent Km value of 0.21 0.04 mm. However, strong negative cooperativity was observed with respect to citrate binding, with a Hill coefficient (nH) of 0.45. Although the dissociation constant of the first citrate molecule was 0.057 0.008 mm, binding of the first citrate molecule to the enzyme drastically decreased the affinity of the enzyme for the second molecule by a factor of 23. ADP was a competitive inhibitor of ATP with a Ki value of 0.037 0.006 mm. Together with previous findings that the enzyme catalyzed the reaction only in the direction of citrate cleavage, these kinetic features indicated that Cl -ACL can regulate both the direction and carbon flux of the RTCA cycle in C. limicola. Furthermore, in order to gain insight on the reaction mechanism, we performed biochemical analyses of Cl -ACL. His273 of the , subunit was indicated to be the phosphorylated residue in the catalytic center, as both catalytic activity and phosphorylation of the enzyme by ATP were abolished in an H273A mutant enzyme. We found that phosphorylation of the subunit was reversible. Nucleotide preference for activity was in good accordance with the preference for phosphorylation of the enzyme. Although residues interacting with nucleotides in the succinyl-CoA synthetase from Escherichia coli were conserved in AclB, AclA alone could be phoshorylated with the same nucleotide specificity observed in the holoenzyme. However, AclB was necessary for enzyme activity and contributed to enhance phosphorylation and stabilization of AclA. [source]

Kinetic evidences for facilitation of peptide channelling by the proteasome activator PA28

FEBS JOURNAL, Issue 20 2000
Ralf Stohwasser
The activation kinetics of constitutive and IFN,-stimulated 20S proteasomes obtained with homomeric (recPA28,, recPA28,) and heteromeric (recPA28,,) forms of recombinant 11S regulator PA28 was analysed by means of kinetic modelling. The activation curves obtained with increasing concentrations of the individual PA28 subunits (RecP28,/RecP28,/RecP28,+ RecP28,) exhibit biphasic characteristics which can be attributed to a low-level activation by PA28 monomers and full proteasome activation by assembled activator complexes. The dissociation constants do not reveal significant differences between the constitutive and the immunoproteasome. Intriguingly, the affinity of the proteasome towards the recPA28,, complex is about two orders of magnitude higher than towards the homomeric PA28, and PA28, complexes. Striking similarities can been revealed in the way how PA28 mediates the kinetics of latent proteasomes with respect to three different fluorogenic peptides probing the chymotrypsin-like, trypsin-like and peptidylglutamyl-peptide hydrolyzing like activity: (a) positive cooperativity disappears as indicated by a lack of sigmoid initial parts of the kinetic curves, (b) substrate affinity is increased, whereby (c), the maximal activity remains virtually constant. As these kinetic features are independent of the peptide substrates, we conclude that PA28 exerts its activating influence on the proteasome by enhancing the uptake (and release) of shorter peptides. [source]

Differential kinetic features by tumour topography in cutaneous small-cell neuroendocrine (Merkel cell) carcinomas

L Pozo
Abstract Background/Objectives Merkel cell carcinomas (MCC) reveal epithelial and neuroendocrine differentiation, but its topographic cell kinetics remains unknown. This study analyses proliferation, apoptosis, and DNA ploidy by topography, features that can help planning therapeutic protocols. This study topographically analyses proliferation, apoptosis, and DNA ploidy. Methods We selected 27 small-cell MCCs (expressing one epithelial and two neural markers, with consistent ultrastructural findings) to evaluate mitotic figure counting, Ki-67 index, apoptosis index based on the in situ end labelling of fragmented DNA (using Escherichia coli DNA polymerase I, Klenow fragment), DNA ploidy, and BCL2 and TP53 immuno-expression. At least 50 high-power fields were screened per topographic compartment (superficial or papillary dermis, and deep or reticular dermis), recording average and standard deviation for each variable. Variables were statistically compared in each tumour compartment using analysis of variance and Student's t -test (significant if P < 0.05). Results MCCs revealed superficial aneuploid DNA content, and no topographic differences for proliferation markers. Apoptosis showed significantly lower values in the deep compartment (average, P = 0.0050, and standard deviation, P = 0.0074), correlating with increased BCL2 and TP53 immuno-expressions. Conclusions High homogeneously distributed proliferation and superficial aneuploid DNA content defines MCCs. Apoptosis follows proliferation in superficial compartments, being less variable and proliferation independent in deep compartments, where it is inversely correlated with BCL2/TP53 expression. [source]

Substrate determines asymmetrical gait dynamics in marmosets (Callithrix jacchus) and squirrel monkeys (Saimiri boliviensis)

Jesse W. Young
Abstract Studies of skeletal pathology indicate that injury from falling accounts for most long bone trauma in free-ranging primates, suggesting that primates should be under strong selection to manifest morphological and behavioral mechanisms that increase stability on arboreal substrates. Although previous studies have identified several kinematic and kinetic features of primate symmetrical gaits that serve to increase arboreal stability, very little work has focused on the dynamics of primate asymmetrical gaits. Nevertheless, asymmetrical gaits typify the rapid locomotion of most primates, particularly in smaller bodied taxa. This study investigated asymmetrical gait dynamics in growing marmosets and squirrel monkeys moving on terrestrial and simulated arboreal supports (i.e., an elevated pole). Results showed that monkeys used several kinematic and kinetic adjustments to increase stability on the pole, including reducing peak vertical forces, limiting center of mass movements, increasing substrate contact durations, and using shorter and more frequent strides (thus limiting disruptive whole-body aerial phases). Marmosets generally showed greater adjustment to pole locomotion than did squirrel monkeys, perhaps as a result of their reduced grasping abilities and retreat from the fine-branch niche. Ontogenetic increases in body size had relatively little independent influence on asymmetrical gait dynamics during pole locomotion, despite biomechanical theory suggesting that arboreal instability is exacerbated as body size increases relative to substrate diameter. Overall, this study shows that 1) symmetrical gaits are not the only stable way to travel arboreally and 2) small-bodied primates utilize specific kinematic and kinetic adjustments to increase stability when using asymmetrical gaits on arboreal substrates. Am J Phys Anthropol, 2009. 2008 Wiley-Liss, Inc. [source]

Oxidation of alkanes and alcohols with hydrogen peroxide catalyzed by complex Os3(CO)10(-H)2,

Georgiy B. Shul'pin
Abstract Trinuclear carbonyl hydride cluster, Os3(CO)10(-H)2, catalyzes oxidation of cyclooctane to cyclooctyl hydroperoxide by hydrogen peroxide in acetonitrile solution. The hydroperoxide partly decomposes in the course of the reaction to afford cyclooctanone and cyclooctanol. Selectivity parameters obtained in oxidations of various linear and branched alkanes as well as kinetic features of the reaction indicated that the alkane oxidation occurs with the participation of hydroxyl radicals. A similar mechanism operates in transformation of benzene into phenol and styrene into benzaldehyde. The system also oxidizes 1-phenylethanol to acetophenone. The kinetic study led to a conclusion that oxidation of alcohols does not involve hydroxyl radicals as main oxidizing species and apparently proceeds with the participation of osmyl species, ,OsO'. Copyright 2010 John Wiley & Sons, Ltd. [source]