Comparison of Two Metal-Dependent Pyruvate Aldolases Related by Convergent Evolution: Substrate Specificity, Kinetic Mechanism, and Substrate Channeling

Biochemistry ◽  
2010 ◽  
Vol 49 (17) ◽  
pp. 3774-3782 ◽  
Author(s):  
Weijun Wang ◽  
Perrin Baker ◽  
Stephen Y. K. Seah
Keyword(s):  
Substrate Specificity ◽  
Convergent Evolution ◽  
Kinetic Mechanism ◽  
Substrate Channeling

scholarly journals Substrate specificity of sheep liver sorbitol dehydrogenase

1998 ◽  
Vol 330 (1) ◽  
pp. 479-487 ◽  
Author(s):  
I. Rune LINDSTAD ◽  
Peter KÖLL ◽  
John S. McKINLEY-McKEE
Keyword(s):  
Substrate Specificity ◽  
Ternary Complex ◽  
Kinetic Mechanism ◽  
Sorbitol Dehydrogenase ◽  
Hydroxyl Group ◽  
Enzyme Active Site ◽  
Sheep Liver ◽  
Rate Limiting Step ◽  
Steady State Kinetics ◽  
Rate Limiting

The substrate specificity of sheep liver sorbitol dehydrogenase has been studied by steady-state kinetics over the range pH 7-10. Sorbitol dehydrogenase stereo-selectively catalyses the reversible NAD-linked oxidation of various polyols and other secondary alcohols into their corresponding ketones. The kinetic constants are given for various novel polyol substrates, including L-glucitol, L-mannitol, L-altritol, D-altritol, D-iditol and eight heptitols, as well as for many aliphatic and aromatic alcohols. The maximum velocities (kcat) and the substrate specificity-constants (kcat/Km) are positively correlated with increasing pH. The enzyme-catalysed reactions occur by a compulsory ordered kinetic mechanism with the coenzyme as the first, or leading, substrate. With many substrates, the rate-limiting step for the overall reaction is the enzyme-NADH product dissociation. However, with several substrates there is a transition to a mechanism with partial rate-limitation at the ternary complex level, especially at low pH. The kinetic data enable the elucidation of new empirical rules for the substrate specificity of sorbitol dehydrogenase. The specificity-constants for polyol oxidation vary as a function of substrate configuration with D-xylo > d-ribo > L-xylo > d-lyxo ≈ l-arabino > D-arabino > l-lyxo. Catalytic activity with a polyol or an aromatic substrate and various 1-deoxy derivatives thereof varies with -CH2OH >-CH2NH2 >-CH2OCH3 ≈-CH3. The presence of a hydroxyl group at each of the remaining chiral centres of a polyol, apart from the reactive C2, is also nonessential for productive ternary complex formation and catalysis. A predominantly nonpolar enzymic epitope appears to constitute an important structural determinant for the substrate specificity of sorbitol dehydrogenase. The existence of two distinct substrate binding regions in the enzyme active site, along with that of the catalytic zinc, is suggested to account for the lack of stereospecificity at C2 in some polyols.

scholarly journals Substrate Specificity and Kinetic Mechanism of Mammalian G9a Histone H3 Methyltransferase

2004 ◽  
Vol 279 (51) ◽  
pp. 53248-53258 ◽  
Author(s):  
Debasis Patnaik ◽  
Hang Gyeong Chin ◽  
Pierre-Olivier Estève ◽  
Jack Benner ◽  
Steven E. Jacobsen ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Histone H3 ◽  
Kinetic Mechanism

scholarly journals Substrate Specificity and Kinetic Mechanism of the Insect Sulfotransferase, Retinol Dehydratase

1998 ◽  
Vol 273 (52) ◽  
pp. 35381-35387 ◽  
Author(s):  
Efsevia Vakiani ◽  
John Gately Luz ◽  
Jochen Buck
Keyword(s):  
Substrate Specificity ◽  
Kinetic Mechanism

scholarly journals Mechanistic Studies of the Yeast Polyamine Oxidase Fms1: Kinetic Mechanism, Substrate Specificity, and pH Dependence

Biochemistry ◽  
2010 ◽  
Vol 49 (49) ◽  
pp. 10440-10448 ◽  
Author(s):  
Mariya S. Adachi ◽  
Jason M. Torres ◽  
Paul F. Fitzpatrick
Keyword(s):  
Substrate Specificity ◽  
Ph Dependence ◽  
Kinetic Mechanism ◽  
Polyamine Oxidase ◽  
Mechanistic Studies

scholarly journals Structure and function of yeast alcohol dehydrogenase

2000 ◽  
Vol 65 (4) ◽  
pp. 207-227 ◽  
Author(s):  
Svetlana Trivic ◽  
Vladimir Leskovac
Keyword(s):  
Alcohol Dehydrogenase ◽  
Substrate Specificity ◽  
Primary Structure ◽  
Active Site ◽  
Hydride Transfer ◽  
Kinetic Mechanism ◽  
Structure And Function ◽  
Chemical Mechanism ◽  
And Function ◽  
Font Color

1. Introduction 2. Isoenzymes of YADH 3. Substrate specificity 4. Kinetic mechanism 5. Primary structure 6. The active site 7. Mutations in the yeast enzyme 8. Chemical mechanism 9. Binding of coenzymes 10. Hydride transfer <br><br><font color="red"><b> This article has been corrected. Link to the correction <u><a href="http://dx.doi.org/10.2298/JSC0008609E">10.2298/JSC0008609E</a><u></b></font>

Kinetic mechanism and substrate specificity of glutathione peroxidase activity of ebselen (PZ51)

1988 ◽  
Vol 37 (11) ◽  
pp. 2267-2271 ◽  
Author(s):  
Matilde Maiorino ◽  
Antonella Roveri ◽  
Mariagrazia Coassin ◽  
Fulvio Ursini
Keyword(s):  
Glutathione Peroxidase ◽  
Substrate Specificity ◽  
Peroxidase Activity ◽  
Kinetic Mechanism ◽  
Glutathione Peroxidase Activity

scholarly journals Pre-steady-state Kinetic Analysis of Amino Acid Transporter SLC6A14 Reveals Rapid Turnover Rate and Substrate Translocation

2021 ◽  
Vol 12 ◽  
Author(s):  
Yueyue Shi ◽  
Jiali Wang ◽  
Elias Ndaru ◽  
Christof Grewer
Keyword(s):  
Steady State ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Turnover Rate ◽  
Family Members ◽  
Kinetic Mechanism ◽  
Amino Acid Transporter ◽  
Slc6 Family ◽  
Acid Transporter ◽  
Rate Limiting

SLC6A14 (solute carrier family 6 member 14) is an amino acid transporter, driven by Na+ and Cl− co-transport, whose structure, function, and molecular and kinetic mechanism have not been well characterized. Its broad substrate selectivity, including neutral and cationic amino acids, differentiates it from other SLC6 family members, and its proposed involvement in nutrient transport in several cancers suggest that it could become an important drug target. In the present study, we investigated SLC6A14 function and its kinetic mechanism after expression in human embryonic kidney (HEK293) cells, including substrate specificity and voltage dependence under various ionic conditions. We applied rapid solution exchange, voltage jumps, and laser photolysis of caged alanine, allowing sub-millisecond temporal resolution, to study SLC6A14 steady state and pre-steady state kinetics. The results highlight the broad substrate specificity and suggest that extracellular chloride enhances substrate transport but is not required for transport. As in other SLC6 family members, Na+ binding to the substrate-free transporter (or conformational changes associated with it) is electrogenic and is likely rate limiting for transporter turnover. Transient current decaying with a time constant of &lt;1ms is also observed after rapid amino acid application, both in forward transport and homoexchange modes, indicating a slightly electrogenic, but fast and not rate-limiting substrate translocation step. Our results, which are consistent with kinetic modeling, suggest rapid transporter turnover rate and substrate translocation with faster kinetics compared with other SLC6 family members. Together, these results provided novel information on the SLC6A14 transport cycle and mechanism, expanding our understanding of SLC6A14 function.

Sign in / Sign up

Export Citation Format

Share Document