Short communicationComputational analysis of di-peptides correlated with the optimal temperature in G/11 xylanase
Introduction
Recently there has been an increasing interest in xylanase (EC 3.2.1.8) for its wide application in industry. It breaks randomly down the internal β-1,4-glycosidic bond of xylan, the major hemicellulose component of the plant cell wall. This makes it a very useful enzyme in animal feed, food and beverage industry, bakery industry, fruit juices-clarifying, and especially in pulp and paper industry [1]. But very often, the biotechnology environment is extreme and demands robust xylanase. For example, pulp bleaching is carried out in a very low pH environment, connected with a hot, caustic treatment of wood, which limits the usage of xylanase. Therefore, thermostable xylanase is of interest [2], [3]. To elevate the optimal temperature of an enzyme, many methods have been employed, such as site-directed mutagenesis (SDM), directed evolution and computational design [4]. To elucidate the thermostability of protein, such methods as crystal structure comparison and genomic comparison have been used [5], [6], [7], [8], [9], and many factors were assumed to be responsible [5], [6], [7], [10]. However, the limited number of known protein structures available in the protein data bank (PDB) makes it difficult to study the thermostability in a single enzyme family; since different proteins use different strategies for maintaining thermostability, the genomic comparison methods may give inconsistent results [11], [12], [13]. Because of the limited number of crystal structures and the prevalence of neutral mutations occurring in proteins, different thermostability analyses often do not agree or even contradict with each other [14]. Enzyme engineering based on these conflicting results often did not lead to an expected conclusion, sometimes even led to the reverse effect [15]. Given the Anfinsen's principle that the fold of a protein is encoded in its amino acid sequence [16], and the large number of protein sequences available in sequence databases, it is necessary to study the relationship between sequence properties and thermostability, we selected xylanase in this study because a large number of sequences of this protein are available in the Swiss-prot databank. Using sequence analysis, we studied the amino acids correlated with the optimal pH in G/11 xylanase [17]; using principle components analysis, we discriminated between these two families [18], and found the di-peptides correlated with the optimal temperature in F/10 xylanase [19].
In the process of analyzing thermostability of enzyme, the optimal growth temperature of the source organism was commonly used [5], [20]. Whereas, it usually cannot reflect the physicochemical property of an enzyme itself, there is often a big difference between the Topt of xylanase and the growth temperature of source organism, sometimes the difference reaches to 33 °C [21]. The xylanase produced by an Aspergillus strain living at 37 °C showed maximal activity at 80 °C [22]. During the construction of the analysis data set, xylanases coming from the same organism were also found to have different Topts; this fact reflects a different thermostability. Based on the investigation of structural similarities, it is commonly held that xylanases have evolved by domain shuffling [23], the study of genomic comparison also indicated that gene transfer occurred between archaea and bacteria [24]. Based on the homologous alignment of G/11 xylanases, we also found that the relationship between sequences is closer than that of the organisms from taxonomic classification. Therefore, it is no advantage to use the optimal growth temperature of source microorganism as criterion, and the Topt of xylanase is used instead in the present study. Because of its low molecular weight, G/11 xylanase is often regarded as an advantageous agent in pulp bleaching compared with that of F/10 xylanase. In the present study, the dipeptides correlated with the optimal temperature of xylanase were computed by using G/11 xylanase sequences from Swiss-Prot (release 43.5.0 of 07-Jun-2004).
Section snippets
Materials and method
Construction of data set. As mentioned above, G/11 xylanase sequences were downloaded from Swiss-Prot (http://www.expasy.org/) (release 43.5 of 07-Jun-2004) for the purpose of its non-redundancy and having been examined by experts, to ensure the accuracy of the sequences under study. There were 32 sequences downloaded from Swiss-Prot. Those G/11 xylanases not deposited in Swiss-prot were not selected as our analysis object, such as the xylanase from Dictyoglomus thermophilum [25], the xylanases
Result and discussion
Seen from Table 1, xylanases from the same organism have different optimal temperature. Such as the xylanases P33557 and P33559 from Aspergillus kawachii, with Topts of 50 and 60 °C, respectively [31], [32]. Although the difference is not as large as to separate them from mesophilic to thermophilic enzymes, it does reflect a different thermostability. If the source microorganism growth temperature were used in the analysis, their real difference would disappear and the stability-affecting
Conclusion
Our results suggest that thermostability is correlated with the order and composition of residues in G/11 xylanase, which was a long controversy between “traffic rule” and genome sequence comparison [42], [43], the controversy might be caused by different proteins used by different researchers, because different protein structures employ different strategy for maintaining thermostability. The results clearly explain the successful improvement of the xylanase thermostability by the introduction
References (43)
- et al.
Purification and biochemical characterization of two xylanases produced by Aspergillus caespitosus and their potential for kraft pulp bleaching
Process Biochem
(2005) - et al.
Production of a xylanolytic enzyme by a thermoalkaliphilic Bacillus sp. JB-99 in solid state fermentation
Process Biochem
(2005) - et al.
Tiny TIM: a small, tetrameric, hyperthermostable triosephosphate isomerase
J Mol Biol
(2001) - et al.
Structural and genomic correlates of hyperthermostability
J Biol Chem
(2000) - et al.
The influence of dipeptide composition on protein thermostability
FEBS Lett
(2004) - et al.
Distributions of structural features contributing to the thermostability in mesophilic and thermophilic ? glycosyl hydrolases
Biochim Biophys Acta
(2000) - et al.
Increasing the thermal stability of cellulase using rules learned from thermophilic proteins: a pilot study
Biophyl Chem
(2002) - et al.
Computational analysis of responsible dipeptides for optimum pH in G/11 xylanase
Biochem Biophys Res Commun
(2004) - et al.
Principle component analysis in F/10 and G/11 xylanase
Biochem Biophys Res Commun
(2004) - et al.
A novel model to calculate dipeptides responsible for optimum temperature in F/10 xylanase
Process Biochem
(2005)
Protein thermal stability, hydrogen bonds, and ion pairs
J Mol Biol
Molecular and biotechnological aspects of xylanases
FEMS Microbiol Rev
Three-dimensional structure of endo-1,4-xylanase I from Aspergillus niger: molecular basis for its low pH optimum
J Mol Biol
A combination of weakly stabilizing mutations with a disulfide in the helix region of Trichoderma reesei endo-1,4-xylanase II increases the thermal stability through synergism
J Biotechnol
The endoxylanases from family 11: computer analysis of protein sequences reveals important structural and phylogenetic relationships
J Biotechnol
Engineering protein thermal stability, sequence statistics point to residues substitutions in alpha-helices
J Mol Biol
Thermostable and alkaline-tolerant microbial cellulase-free xylanases produced from agricultural wastes and the properties required for use in pulp bleaching bioprocesses: a review
Process Biochem
Hyperthermophilic enzymes: sources, uses, and molecular mechanisms for thermostability
Microbio Mol Bio Rev
High resolution structure and sequence of T. aurantiacus xylanase. I. Implication for the evolution of thermostability in family 10 xylanases and enzymes with Barrel architecture
Proteins
Three-dimensional structures of thermophilic beta-1,4-xylanases from Chaetomium thermophilum and Nonomuraea flexuosa—comparison of twelve xylanases in relation to their thermal stability
Eur J Biochem
Identification of thermophilic species by the amino acid compositions deduced from their genomes
Nucl Acid Res
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