Crystal structure determination and inhibition studies of a novel xylanase and alpha-amylase inhibitor protein (XAIP) from Scadoxus multiflorus.
A novel plant protein isolated from the underground bulbs of Scadoxus multiflorus, xylanase and alpha-amylase inhibitor protein (XAIP), inhibits two structurally and functionally unrelated enzymes: xylanase and alpha-amylase. The mature protein contains 272 amino acid residues which show sequence identities of 48% to the plant chitinase hevamine and 36% to xylanase inhibitor protein-I, a double-headed inhibitor of GH10 and GH11 xylanases. However, unlike hevamine, it is enzymatically inactive and, unlike xylanase inhibitor protein-I, it inhibits two functionally different classes of enzyme. The crystal structure of XAIP has been determined at 2.0 A resolution and refined to R(cryst) and R(free) factors of 15.2% and 18.6%, respectively. The polypeptide chain of XAIP adopts a modified triosephosphate isomerase barrel fold with eight beta-strands in the inner circle and nine alpha-helices forming the outer ring. The structure contains three cis peptide bonds: Gly33-Phe34, Tyr159-Pro160 and Trp253-Asp254. Although hevamine has a long accessible carbohydrate-binding channel, in XAIP this channel is almost completely filled with the side-chains of residues Phe13, Pro77, Lys78 and Trp253. Solution studies indicate that XAIP inhibits GH11 family xylanases and GH13 family alpha-amylases through two independent binding sites located on opposite surfaces of the protein. Comparison of the structure of XAIP with that of xylanase inhibitor protein-I, and docking studies, suggest that loops alpha3-beta4 and alpha4-beta5 may be involved in the binding of GH11 xylanase, and that helix alpha7 and loop beta6-alpha6 are suitable for the interaction with alpha-amylase.
Kumar
Sanjit
S
Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
Singh
Nagendra
N
Sinha
Mau
M
Dube
Divya
D
Singh
S Baskar
SB
Bhushan
Asha
A
Kaur
Punit
P
Srinivasan
Alagiri
A
Sharma
Sujata
S
Singh
Tej P
TP
D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D043364:Endo-1,4-beta Xylanases / Q000037:antagonists & inhibitors*; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D018517:Plant Roots / Q000737:chemistry; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
D004791:Enzyme Inhibitors; D010940:Plant Proteins; D000516:alpha-Amylases; D043364:Endo-1,4-beta Xylanases