scholarly journals Structures of lipoyl synthase reveal a compact active site for controlling sequential sulfur insertion reactions

2014 ◽  
Vol 464 (1) ◽  
pp. 123-133 ◽  
Author(s):  
Jenny E. Harmer ◽  
Martyn J. Hiscox ◽  
Pedro C. Dinis ◽  
Stephen J. Fox ◽  
Andreas Iliopoulos ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
Binding Sites ◽  
Substrate Binding ◽  
Insertion Reactions ◽  
Substrate Binding Sites ◽  
Lipoyl Synthase

The biosynthesis of lipoyl cofactors requires two lipoyl synthase-mediated sulfur insertions. We report the crystal structures of a lipoyl synthase complexed with S-adenosylhomocysteine or 5′-methylthioadenosine. Models based on these structures identify likely substrate-binding sites.

Crystal structures of the GH6 Orpinomyces sp. Y102 CelC7 enzyme with exo and endo activity and its complex with cellobiose

2019 ◽  
Vol 75 (12) ◽  
pp. 1138-1147
Author(s):  
Hsiao-Chuan Huang ◽  
Liu-Hong Qi ◽  
Yo-Chia Chen ◽  
Li-Chu Tsai
Keyword(s):  
Enzymatic Hydrolysis ◽  
Crystal Structures ◽  
Active Site ◽  
Binding Sites ◽  
Glycoside Hydrolase ◽  
Catalytic Domain ◽  
Glycoside Hydrolase Family ◽  
Substrate Binding Sites ◽  
Key Residues ◽  
Hydrolase Family

The catalytic domain (residues 128–449) of the Orpinomyces sp. Y102 CelC7 enzyme (Orp CelC7) exhibits cellobiohydrolase and cellotriohydrolase activities. Crystal structures of Orp CelC7 and its cellobiose-bound complex have been solved at resolutions of 1.80 and 2.78 Å, respectively. Cellobiose occupies subsites +1 and +2 within the active site of Orp CelC7 and forms hydrogen bonds to two key residues: Asp248 and Asp409. Furthermore, its substrate-binding sites have both tunnel-like and open-cleft conformations, suggesting that the glycoside hydrolase family 6 (GH6) Orp CelC7 enzyme may perform enzymatic hydrolysis in the same way as endoglucanases and cellobiohydrolases. LC-MS/MS analysis revealed cellobiose (major) and cellotriose (minor) to be the respective products of endo and exo activity of the GH6 Orp CelC7.

Papain-like lysosomal cysteine proteases and their inhibitors: drug discovery targets?

2003 ◽  
Vol 70 ◽  
pp. 15-30 ◽  
Author(s):  
Dŭsan Turk ◽  
Boris Turk ◽  
Vito Turk
Keyword(s):  
Drug Discovery ◽  
Active Site ◽  
Binding Sites ◽  
Substrate Binding ◽  
Cysteine Proteases ◽  
Structural Features ◽  
Discovery Research ◽  
Drug Discovery Research ◽  
Lysosomal Cysteine Proteases ◽  
Substrate Binding Sites

Papain-like lysosomal cysteine proteases are processive and digestive enzymes that are expressed in organisms from bacteria to humans. Increasing knowledge about the physiological and pathological roles of cysteine proteases is bringing them into the focus of drug discovery research. These proteases have rather short active-site clefts, comprising three well defined substrate-binding subsites (S2, S1 and S1') and additional broad binding areas (S4, S3, S2' and S3'). The geometry of the active site distinguishes cysteine proteases from other protease classes, such as serine and aspartic proteases, which have six and eight substrate-binding sites respectively. Exopeptidases (cathepsins B, C, H and X), in contrast with endopeptidases (such as cathepsins L, S, V and F), possess structural features that facilitate the binding of N- and C-terminal groups of substrates into the active-site cleft. Other than a clear preference for free chain termini in the case of exopeptidases, the substrate-binding sites exhibit no strict specificities. Instead, their subsite preferences arise more from the specific exclusion of substrate types. This presents a challenge for the design of inhibitors to target a specific cathepsin: only the cumulative effect of an assembly of inhibitor fragments will bring the desired result.

Crystal Structures ofPasteurella multocidaSialyltransferase Complexes with Acceptor and Donor Analogues Reveal Substrate Binding Sites and Catalytic Mechanism†,‡

Biochemistry ◽  
2007 ◽  
Vol 46 (21) ◽  
pp. 6288-6298 ◽  
Author(s):  
Lisheng Ni ◽  
Harshal A. Chokhawala ◽  
Hongzhi Cao ◽  
Ryan Henning ◽  
Laura Ng ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Binding Sites ◽  
Catalytic Mechanism ◽  
Substrate Binding ◽  
Substrate Binding Sites

scholarly journals Sigmoidal kinetic model for two co-operative substrate-binding sites in a cytochrome P450 3A4 active site: an example of the metabolism of diazepam and its derivatives

1999 ◽  
Vol 340 (3) ◽  
pp. 845-853 ◽  
Author(s):  
Magang SHOU ◽  
Qin MEI ◽  
JR. Michael W. ETTORE ◽  
Renke DAI ◽  
Thomas A. BAILLIE ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Kinetic Model ◽  
Binding Affinity ◽  
Active Site ◽  
Binding Sites ◽  
Dissociation Constants ◽  
Substrate Binding ◽  
Cytochrome P450 3A4 ◽  
Multiple Drugs ◽  
Substrate Binding Sites

Cytochrome P450 3A4 (CYP3A4) plays a prominent role in the metabolism of a vast array of drugs and xenobiotics and exhibits broad substrate specificities. Most cytochrome P450-mediated reactions follow simple Michaelis-Menten kinetics. These parameters are widely accepted to predict pharmacokinetic and pharmacodynamic consequences in vivo caused by exposure to one or multiple drugs. However, CYP3A4 in many cases exhibits allosteric (sigmoidal) characteristics that make the Michaelis constants difficult to estimate. In the present study, diazepam, temazepam and nordiazepam were employed as substrates of CYP3A4 to propose a kinetic model. The model hypothesized that CYP3A4 contains two substrate-binding sites in a single active site that are both distinct and co-operative, and the resulting velocity equation had a good fit with the sigmoidal kinetic observations. Therefore, four pairs of the kinetic estimates (KS1, kα, KS2, kβ, KS3, k∆, KS4 and kγ) were resolved to interpret the features of binding affinity and catalytic ability of CYP3A4. Dissociation constants KS1 and KS2 for two single-substrate-bound enzyme molecules (SE and ES) were 3-50-fold greater than KS3 and KS4 for a two-substrate-bound enzyme (SES), while respective rate constants k∆ and kγ were 3-218-fold greater than kα and kβ, implying that access and binding of the first molecule to either site in an active pocket of CYP3A4 can enhance the binding affinity and reaction rate of the vacant site for the second substrate. Thus our results provide some new insights into the co-operative binding of two substrates in the inner portions of an allosteric CYP3A4 active site.

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