The role of disulfide bonds in the conformational stability and catalytic activity of phytase

2004 ◽  
Vol 82 (2) ◽  
pp. 329-334 ◽  
Author(s):  
Xiao-Yun Wang ◽  
Fan-Guo Meng ◽  
Hai-Meng Zhou
Keyword(s):  
Catalytic Activity ◽  
Disulfide Bonds ◽  
Conformational Stability ◽  
Three Dimensional ◽  
Fluorescence Emission ◽  
Dimensional Structure ◽  
Enzyme Activity Assay ◽  
First Order ◽  
Far Ultraviolet

Previous studies have predicted five disulfide bonds in Aspergillus niger phytase (phy A). To investigate the role of disulfide bonds, intrinsic fluorescence spectra, far-ultraviolet circular dichroism (CD) spectra, and an enzyme activity assay were used to compare the differences of catalytic activity and conformational stability of phytase during denaturation in urea in the presence and absence of dithiothreitol (DTT). In the presence of 2 mM DTT, the inactivation and unfolding were greatly enhanced at the same concentration of denaturant. The fluorescence emission maximum red shift and decreases of ellipticity at 222 nm were in accord with the changes of catalytic activity. The kinetics of the unfolding courses were a biphasic process consisting of two first-order reactions in the absence of DTT and a monophasic process of a first-order reaction in the presence of DTT. The results suggested that the loss of enzymatic activity was most likely because of a conformational change, and that disulfide bonds played an important role in three-dimensional structure and catalytic activity.Key words: phytase, urea denaturation, inactivation, disulfide bond.

scholarly journals Role of Simian Virus 40 Vp1 Cysteines in Virion Infectivity

2000 ◽  
Vol 74 (23) ◽  
pp. 11388-11393 ◽  
Author(s):  
Peggy P. Li ◽  
Akira Nakanishi ◽  
Mary A. Tran ◽  
Adler M. Salazar ◽  
Robert C. Liddington ◽  
...  
Keyword(s):  
Life Cycle ◽  
Disulfide Bonds ◽  
Three Dimensional ◽  
Simian Virus 40 ◽  
Experimental System ◽  
Simian Virus ◽  
Dimensional Structure ◽  
Wild Type

ABSTRACT We have developed a new nonoverlapping infectious viral genome (NO-SV40) in order to facilitate structure-based analysis of the simian virus 40 (SV40) life cycle. We first tested the role of cysteine residues in the formation of infectious virions by individually mutating the seven cysteines in the major capsid protein, Vp1. All seven cysteine mutants—C9A, C49A, C87A, C104A, C207S, C254A, and C267L—retained viability. In the crystal structure of SV40, disulfide bridges are formed between certain Cys104 residues on neighboring pentamers. However, our results show that none of these disulfide bonds are required for virion infectivity in culture. We also introduced five different mutations into Cys254, the most strictly conserved cysteine across the polyomavirus family. We found that C254L, C254S, C254G, C254Q, and C254R mutants all showed greatly reduced (around 100,000-fold) plaque-forming ability. These mutants had no apparent defect in viral DNA replication. Mutant Vp1's, as well as wild-type Vp2/3, were mostly localized in the nucleus. Further analysis of the C254L mutant revealed that the mutant Vp1 was able to form pentamers in vitro. DNase I-resistant virion-like particles were present in NO-SV40-C254L-transfected cell lysate, but at about 1/18 the amount in wild-type-transfected lysate. An examination of the three-dimensional structure reveals that Cys254 is buried near the surface of Vp1, so that it cannot form disulfide bonds, and is not involved in intrapentamer interactions, consistent with the normal pentamer formation by the C254L mutant. It is, however, located at a critical junction between three pentamers, on a conserved loop (G2H) that packs against the dual interpentamer Ca2+-binding sites and the invading C-terminal helix of an adjacent pentamer. The substitution by the larger side chains is predicted to cause a localized shift in the G2H loop, which may disrupt Ca2+ ion coordination and the packing of the invading helix, consistent with the defect in virion assembly. Our experimental system thus allows dissection of structure-function relationships during the distinct steps of the SV40 life cycle.

scholarly journals Analysis of structural similarities between brain Thy-1 antigen and immunoglobulin domains. Evidence for an evolutionary relationship and a hypothesis for its functional significance

1981 ◽  
Vol 195 (1) ◽  
pp. 31-40 ◽  
Author(s):  
F E Cohen ◽  
J Novotný ◽  
M J E Sternberg ◽  
D G Campbell ◽  
A F Williams
Keyword(s):  
Functional Significance ◽  
Structure Prediction ◽  
Secondary Structure Prediction ◽  
Three Dimensional ◽  
Evolutionary Relationship ◽  
Dimensional Structure ◽  
Sequence Homologies ◽  
Beta Structure ◽  
Beta 2 Microglobulin

The Thy-1 membrane glycoprotein from rat brain is shown to have structural and sequence homologies with immunoglobulin (Ig) domains on the basis of the following evidence. 1. The two disulphide bonds of Thy-1 are both consistent with the Ig-fold. 2. The molecule contains extensive beta-structure as shown by the c.d. spectrum. 3. Secondary structure prediction locates beta-strands along the sequence in a manner consistent with the Ig-fold. 4. On the basis of rules derived from known beta-sheet structures, a three-dimensional structure with the Ig-fold is predicted as favourable for Thy-1. 5. Sequences in the proposed beta-strands of Thy-1 and known beta-strands of Ig domains show significant sequence homology. This homology is statistically more significant than for the comparison of proposed beta-strand sequences of beta 2-microglobulin with Ig domains. An hypothesis is presented for the possible functional significance of an evolutionary relationship between Thy-1 and Ig. It is suggested that both Thy-1 and Ig evolved from primitive molecules, with an Ig fold, which mediated cell--cell interactions. The present-day role of Thy-1 may be similar to that of the primitive domain.

3D Structure of Striations in the North Pacific

2021 ◽  
Author(s):  
YU ZHANG ◽  
YU PING GUAN ◽  
RUI XIN HUANG
Keyword(s):  
North Pacific ◽  
3D Structure ◽  
Three Dimensional ◽  
Vertical Plane ◽  
Dimensional Structure ◽  
The North ◽  
Layer Analysis ◽  
Fluid Potential ◽  
The North Pacific

AbstractOcean striations are composed of alternating quasi-zonal band-like flows; this kind of organized structure of currents be found in all world’s oceans and seas. Previous studies have mainly been focused on the mechanisms of their generation and propagation. This study uses the spatial high-pass filtering to obtain the three-dimensional structure of ocean striations in the North Pacific in both the z-coordinate and σ-coordinate based on 10-yr averaged SODA3 data. First, we identify an ideal-fluid potential density domain where the striations are undisturbed by the surface forcing and boundary effects. Second, using the isopycnal layer analysis, we show that on isopycnal surfaces the orientations of striations nearly follow the potential vorticity (PV) contours, while in the meridional-vertical plane the central positions of striations are generally aligned with the latitude of zero gradient of the relative PV. Our analysis provides a simple dynamical interpretation and better understanding for the role of ocean striations.

Overall Structure Of The Heparin Molecule

1981 ◽  
Author(s):  
Edward Atkins
Keyword(s):  
Model Building ◽  
Specific Binding ◽  
Three Dimensional ◽  
Local Variation ◽  
Coagulation Cascade ◽  
Dimensional Structure ◽  
Chemical Analyses ◽  
X Ray Diffraction ◽  
First Order ◽  
Iduronic Acid

The polysaccharide suppresses the coagulation of blood by controlling the rate at which the plasma protein antithrombin inactivates the proteases of the coagulation cascade. One facet necessary in the full understanding of the interaction of heparin with antithrombin is the detailed shape of these macromolecules at an atomic resolution. X-ray diffraction methods are necessary to obtain the required information since the wavelength of the radiation is comparable with the distances between atoms. Analysis of the diffraction signals coupled with computerised model building and energy minimization procedures enables the three dimensional structure, or conformation, of the heparin molecule to be examined. A major part of the molecule is a repeating disaccharide of alternating 1.4-linked-2-deoxy-2-sulphamino-α-D-glucose-6-sulphate and 1.4-linked 2-sulphate-α-L-idopyranoslyluronic acid. This is a first order statement, made necessary to avoid confused and sometimes wrongful interpretations of past chemical analyses (which for many years suggested β-D-gluco-pyranosyluronic acid on the prominent uronic acid in heparin). More recent chemical analyses of selected heparins exhibiting high anticoagulent activity indicates that the idealised polydisaccharide repeat is perturned by local variation in the chemistry. The absence of a sulphate appendage on iduronic acid and the replacement of iduronic acid by glucuronic acid in the near vicinity are features proposed. The effect of these chemical modifications on the local conformation of the heparin molecule is considerable and suggest a specific binding site region within the heparin macromolecule amenable to interaction with antithrombin.

scholarly journals The three-dimensional structure of a class-Pi glutathione S-transferase complexed with glutathione: the active-site hydration provides insights into the reaction mechanism

1998 ◽  
Vol 333 (3) ◽  
pp. 811-816 ◽  
Author(s):  
Antonio PÁRRAGA ◽  
Isabel GARCÍA-SÁEZ ◽  
Sinead B. WALSH ◽  
Timothy J. MANTLE ◽  
Miquel COLL
Keyword(s):  
Conformational Changes ◽  
Active Site ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Water Molecules ◽  
X Ray Diffraction ◽  
Glutathione S Transferase ◽  
X Ray ◽  
The Right

The structure of mouse liver glutathione S-transferase P1-1 complexed with its substrate glutathione (GSH) has been determined by X-ray diffraction analysis. No conformational changes in the glutathione moiety or in the protein, other than small adjustments of some side chains, are observed when compared with glutathione adduct complexes. Our structure confirms that the role of Tyr-7 is to stabilize the thiolate by hydrogen bonding and to position it in the right orientation. A comparison of the enzyme–GSH structure reported here with previously described structures reveals rearrangements in a well-defined network of water molecules in the active site. One of these water molecules (W0), identified in the unliganded enzyme (carboxymethylated at Cys-47), is displaced by the binding of GSH, and a further water molecule (W4) is displaced following the binding of the electrophilic substrate and the formation of the glutathione conjugate. The possibility that one of these water molecules participates in the proton abstraction from the glutathione thiol is discussed.

Role of LHCII-containing macrodomains in the structure, function and dynamics of grana

2000 ◽  
Vol 27 (3) ◽  
pp. 279 ◽  
Author(s):  
G. Garab ◽  
L. Mustárdy
Keyword(s):  
Excitation Energy ◽  
Long Range ◽  
Structural Changes ◽  
Higher Plants ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Lateral Heterogeneity ◽  
Long Distance ◽  
3D Network

In higher plants and green algae two types of thylakoids are distinguished, granum (stacked) and stroma (unstacked) thylakoids. They form a three-dimensional (3D) network with large lateral heterogeneity: photosystem II (PSII) and the associated main chlorophyll a/b light-harvesting complex (LHCII) are found predominantly in the stacked region, while PSI and LHCI are located mainly in the unstacked region of the membrane. This picture emerged from the discovery of the physical separation of the two photosystems (Boardman and Anderson 1964). Granal chloroplasts possess significant flexibility, which is essential for optimizing the photosynthetic machinery under various environmental conditions. However, our understanding concerning the assembly, structural dynamics and regulatory functions of grana is far from being complete. In this paper we overview the significance of the three-dimensional structure of grana in the absorption properties, ionic equilibrations, and in the diffusion of membrane components between the stacked and unstacked regions. Further, we discuss the role of chiral macrodomains in the grana. Lateral heterogeneity of thylakoid membranes is proposed to be a consequence of the formation of macrodomains constituted of LHCII and PSII; their long range order permits long distance migration of excitation energy, which explains the energetic connectivity of PSII particles. The ability of macrodomains to undergo light-induced reversible structural changes lends structural flexibility to the granum. In purified LHCII, which has also been shown to form stacked lamellar aggregates with long range chiral order, excitation energy migrates for large distances; these macroaggregates are also capable of undergoing light-induced reversible structural changes and fluorescence quenching. Hence, some basic properties of grana appear to originate from its main constituent, the LHCII.

scholarly journals Role of Computational Methods in Going beyond X-ray Crystallography to Explore Protein Structure and Dynamics

2018 ◽  
Vol 19 (11) ◽  
pp. 3401 ◽  
Author(s):  
Ashutosh Srivastava ◽  
Tetsuro Nagai ◽  
Arpita Srivastava ◽  
Osamu Miyashita ◽  
Florence Tama
Keyword(s):  
Protein Dynamics ◽  
Computational Methods ◽  
Protein Structures ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Insight Into

Protein structural biology came a long way since the determination of the first three-dimensional structure of myoglobin about six decades ago. Across this period, X-ray crystallography was the most important experimental method for gaining atomic-resolution insight into protein structures. However, as the role of dynamics gained importance in the function of proteins, the limitations of X-ray crystallography in not being able to capture dynamics came to the forefront. Computational methods proved to be immensely successful in understanding protein dynamics in solution, and they continue to improve in terms of both the scale and the types of systems that can be studied. In this review, we briefly discuss the limitations of X-ray crystallography in studying protein dynamics, and then provide an overview of different computational methods that are instrumental in understanding the dynamics of proteins and biomacromolecular complexes.

Sign in / Sign up

Export Citation Format

Share Document