High resolution structure and sequence ofT. aurantiacus Xylanase I: Implications for the evolution of thermostability in family 10 xylanases and enzymes with ??-barrel architecture

Abstract

Xylanase I is a thermostable xylanase from the fungus Thermoascus aurantiacus, which belongs to family 10 in the current classification of glycosyl hydrolases. We have determined the three-dimensional X-ray structure of this enzyme to near atomic resolution (1.14 A) by molecular replacement, and thereby corrected the chemically determined sequence previously published. Among the five members of family 10 enzymes for which the structure has been determined, Xylanase I from T. aurantiacus and Xylanase Z from C. thermocellum are from thermophilic organisms. A comparison with the three other available structures of the family 10 xylanases from mesophilic organisms suggests that thermostability is effected mainly by improvement of the hydrophobic packing, favorable interactions of charged side chains with the helix dipoles and introduction of prolines at the N-terminus of helices. In contrast to other classes of proteins, there is very little evidence for a contribution of salt bridges to thermostability in the family 10 xylanases from thermophiles. Further analysis of the structures of other proteins from thermophiles with eight-fold (beta)alpha-barrel architecture suggests that favorable interactions of charged side chains with the helix dipoles may be a common way in which thermophilic proteins with this fold are stabilized. As this is the most common type of protein architecture, this finding may provide a useful guide for site-directed mutagenesis aimed to improve the thermostability of (beta)alpha-barrel proteins. Proteins 1999;36:295-306.

EMERGING ROLE OF N- AND C-Terminal INTERACTIONS IN STABILIZING (Β;/Α) 8 FOLD WITH SPECIAL EMPHASIS ON FAMILY 10 XYLANASES

“…(51, 52), Bacillus stearothermophilus T-6 (53), Bacillus sp . NG-27(8), Bacillus sp. N16-5 (54), Bacillus halodurans (55), Bacillus firmus (56), Caldicellulosiruptor sp .…”

“…N16-5 (54), Bacillus halodurans (55), Bacillus firmus (56), Caldicellulosiruptor sp . (57), Clostridium thermocellum (58), Rhodothermus marinus (59) and Thermoascus aurantiacus (8, 60). A family 10 xylanase isolated from Thermotoga sp .…”

“…It also helps in understanding the underlying principle of protein structure-function relationships providing insights into the thermal stability of GH10 xylanases. A comparison of family 10 xylanase isolated from thermophilic ( Thermoascus aurantiacus and C. thermocellum ) and mesophilic sources highlighted the role of hydrophobic packing, interaction of helix dipole with charged side chains and increase in proline content at the N-terminal of helices in enzyme thermostability (8). The stability and catalytic activity of an enzyme are also affected by metal ions.…”

“…Mutational studies involving T4 lysozyme and barnase have shown that protein stability is highly influenced by the nature and the position of the mutation where it was introduced (6, 7). Structural studies have shown that each class of enzymes has evolved specific adaptation strategies against extreme conditions and structural differences between the families are the basis for this difference in adaptation strategies (8, 2, 9). As a corollary, it is logical to say that each class of proteins have evolved its own mechanism to enhance protein stability under extreme conditions rather than converging on a single universal mechanism and hence necessary to identify the determinants of protein stability for each class of proteins.…”

Xylanase (GH11) from Acremonium cellulolyticus: homologous expression and characterization

“…GH10 xylanases generally have a molecular weight ≥30 kDa and a low p I , while GH11 xylanases are generally smaller (approximately 20 kDa) and have a high p I (Beaugrand et al [2004]). The crystal structures of xylanases show that GH10 enzymes fold into a (β/α) 8 -barrel (Dimarogona et al [2012]; Harris et al [1994]; Lo Leggio et al [1999]), whereas family 11 enzymes have a β-jellyroll structure (Paës et al [2012]; Sidhu et al [1999]). Cellulose and xylan are closely linked together in plant cell walls (Carpita and Gibeaut [1993]).…”