1. Significant observations
Towards Understanding of the Heterotetrameric Structure of ADP-Glucose Pyrophosphorylase (AGPase) Enzyme in Wheat
Ritu Batraa, Saripalli Gautamb, HS Balyana,b, PK Guptaa,b, Kulvinder S Gillc
aBioinformatics Infrastructure Facility, Department of Genetics and Plant Breeding, Charan Singh University, Meerut
bMolecular Biology laboratory, Department of Genetics and Plant Breeding, Ch.Charan Singh University, Meerut
 cDepartment of Crops and Soil Sciences, Washington State University, Pullman, USA.
Introduction
Table 3: Comparative quality analysis of 3D structures of wheat AGPase LS and SS
In the absence of experimental 3D structures, comparative modeling of protein is considered as one of the most accurate method of model building and to understand the function. In the present study, we generated 3D structures of wheat AGPase LS and SS through Swiss-Model server which will be used to get an insight into heterotetrameric structure of wheat AGPase.
Swiss-Model
SAVES
PROCHEK
AGPase
Modelling rogram
Dfire energy
QMEANscore
Z-score
ERRAT
(Quality Factor)
3D -1D (%)
(Verify3D)
Most favoured
region (%) LS
Phyre2
0.675
-1.090
64.25
93.12
82.1
Swiss Model
0.695
-0.859
76.01
97.0
90.4
I-TESSER
0.619
-1.774
91.4
94.9
75.3 SS
0.678
-1.049
90.21
96.81
80.0
0.715
-0.415
82.78
99.32
0.655
-1.322
94.37
91.56
76.60
Methodology followed
Step1
Protein sequence retrieval of wheat AGPase subunits using well characterized maize AGPase genes Sh2 (encoding LS) and Bt2 (encoding SS) from NCBI.
Step2
Identification of conserved domains (CDs) in the reference sequences using Conserved domain database at NCBI (
Step3
Analysis of the protein sequences showing maximum coverage (97%, 79%), maximum percent identity (79%, 90%) and the same conserved domain as present in the reference species maize.
Results
Figure 2: Structures of wheat AGPase LS and SS using potato SS homotetramer (Jin et al. 2005) (PDB ID: 1Yp4.1B) as a template by Swiss-Model server having identity of % and 90.81% respectively with the template.
Significant observations
Two major CD’s were found each in both LS as well as SS, which were similar.
Swiss-Model was found to be the most appropriate model for generating 3D structures of wheat AGPase LS and SS.
Graphical data for wheat AGPase LS and SS for Anolea, Qmean and GROMOS showed negative energy values for both the subunits of AGPase suggesting a favourable energy environment for given amino acids indicating that the 3D models of AGPase LS and SS generated by Swiss-Model are appropriate for generating heterotetrameric structure of AGPase in wheat (data not presented).
Figure 1: Conserved domains in wheat AGPase LS and SS
Table 1: Results of primary sequence analysis of wheat AGPase LS and SS
AGPase
I.D.
Length
Mol wt. (Da) pI -R LS
ABG88200
522 aa
6.12 65 SS
AAM10977 
473 aa
5.53 61 +R
E.C.
I.I.
A.I.
GRAVY 60
45435
42.86
80.52
-0.253 49
46550
39.98
90.97
-0.214
Future perspectives
This study will be helpful to identify crucial amino acids, which may be targeted in future wet lab efforts to develop thermostable AGPase variants in wheat.
References
Jin XS, Ballicora MA, Preiss J, Geiger JH (2005) Crystal structure of potato tuber ADP-glucose pyrophosphorylase. EMBO J 24:694–704
Table 2: Results of secondary sequence analysis of wheat AGPase LS and SS
Acknowledgements
AGPase
α-helix
(%)
Extended Strand
β-Turn
Random Coil LS
30.27
21.84
11.11
36.78 SS
26.85
23.89
12.26
37.00
Authors are thankful to Head, Department of Genetics and Plant Breeding for providing the facilities. RB and SG were awarded Research Associateship and Senior Research Fellowship by Department of Biotechnology, New Delhi, India and PKG and HSB were awarded position of Senior Scientist by Indian National Science Academy, New Delhi during the tenure of this work.