## 4 Conclusions  
In summary, in this paper, the modeled structures of the catalytic domains of bacterial GH17 transglycos from _P. aeruginosa (Glt1)_, _P. putida (Glt3)_ and _B. diazoefficients (Glt20)_ were analyzed, and laminaripentaose and laminarihexaose were docked in the catalytic clefts in order to predict the potential interaction between ligands and enzyme. Results from this computational study, combined with previously reported experimental evidence allowed elaborating on a mechanism for the transfer-reaction of both non-reducing end and reducing-end acting transglycos, based on binding of the donor substrates in the same orientation in both types of enzymes, while loops in the \(-4\) subsites differ. Despite release from either reducing or non-reducing end, subsites \(+1\) and \(+2\) appear to be most conserved and the difference in release is likely due to changes in hydrogen bonding. Substrate docking in Glt20 indicate that presence of covalently bound donor creates space to accommodate the acceptor oligosaccharide in alternative subsites in the catalytic cleft, promoting a branching point and formation of a 1,6-linkage, and this may be due to the conformation of the \(\alpha\)8-helix, which adopts different conformations in every protein studied. The minimum donor size of DP5, can be explained assuming preferred binding of DP4 substrates in subsite \(-4\) to \(-1\), preventing catalysis.  
The following are available online at [[https://www.mdpi.com/article/10.3390/app11094048/s1](https://www.mdpi.com/article/10.3390/app11094048/s1)]([https://www.mdpi.com/article/10.3390/app11094048/s1](https://www.mdpi.com/article/10.3390/app11094048/s1)), Table S1: Parameters used for the prediction of the three-dimensional  
Figure 8: Complexes of Glt20 with different ligands. The hydrophobicity surfaces represent the polar residues in blue and the non-polar in red. White represent areas of intermediate polarity. Subsite amino acids: in orange (\(-3\)), cyan (\(-2\)), purple (\(-1\)), green (\(+1\)) and yellow (\(+2\)) (**A**) Complex Glt20-laminaripentaose (ligand in green). (**B**) Covalent intermediate Glt20-laminaritirose (in green), the covalent bond between sugar and enzyme is indicated by the black arrow. (**C**) Complex Glt20-laminaritirose (green) covalently bound and with docked laminaripentaose acceptor (in pink). Branch subsites are colored in dark pink (\(-3\)* and \(-2\)*).  
structure of _Glls_, Table S2: Contribution of fragments from other models to the hybrid models, Table S3: Mutagenic primers, Figure S1: Activity analysis of the \(wt\) MalE-_Gll20_ and the two single mutants of MalE-_Gll20_:E120Q_ and _E209Q_.  
[PERSON], [PERSON] and [PERSON] designed the current study. [PERSON], [PERSON] and [PERSON] designed the constructs and did different parts of the cloning production and purification of the enzymes. [PERSON] executed the modeling and molecular dynamics of the catalytic modules. All authors took part in analysis of the results and writing of the manuscript. All authors have read and agreed to the published version of the manuscript.  
This research was funded by the Icelandic Centre of Research (grant number 141341-053), the Swedish Research Council Formas (MariKat, grant No. 2019-02359) and by Macro cascade, a project that has received funding from the Bio-Based Industries Joint Undertaking under the European Union Horizon 2020 research and innovation program under grant agreement No. 720755.  
Not applicable.  
The files of the models presented in this study are available on request from the corresponding author. Other data are available in the article or the Supplementary Materials.  
The computational simulations were enabled by resources provided by the Swedish National Infrastructure for Computing (SNIC) at Lunarc at Lund University, partially funded by the Swedish Research Council through grant agreement No. 2018-05973.  
The authors declare no conflict of interest.