Homology Modelling of the b Fraction of Factor B

Faller, Mark G (1997) Homology Modelling of the b Fraction of Factor B. MSc(R) thesis, University of Glasgow.

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The b domain of factor B (Bb) is found to be moderately homologous to the mammalian serine proteases. Sequence homology analysis showed that out of all the known serine protease structures, Bb was most homologous to bovine trypsin. Sequence alignment between Bb and bovine trypsin gave a sequence identity of the two sequences of between 17 - 21%. This meant that protein model building of Bb from a known serine protease structure, in this case bovine trypsin, could be attempted. The structure of bovine trypsin was taken from the Brookhaven Database. All the molecular modelling was carried out with the software package "COMMET" that was developed in our laboratory. In the first instance all of the substitutions necessary to computationally mutate the bovine trypsin structure into the Bb structure were carried out. Substitutions were carried out first as they are the least disruptive of the three modelling techniques used in homology modelling. Substitution only alters the side chain atoms of the residue that is being modified. No alterations to the backbone atoms are necessary at this stage. The software keeps the new side chain position as close to the original as possible. Where this is not feasible the side chain conformation is determined by a conformational search of the side chain's conformation space. The deletions from bovine trypsin were all small and accomplished by simple removal of the appropriate residues followed by energy minimisation to reposition and rejoin the main chain. Small insertions up to three residues long were built using the "insert" routine. After inserting a residue its side chain's torsion angles were defined by a conformational search. To ease steric strain at the site of small deletions and insertion a segment five residues either side of the insertion or deletion was run through the energy minimiser. There are a total of eight insertions of three residues in length or longer: Gin 30: 3 residues in length Ser 186: 3 residues in length Gly 129: 7 residues in length His 231: 8 residues in length Glu 101: 9 residues in length Leu 143: 9 residues in length Arg 170: 13 residues in length The conformation of these large insertions was calculated using a sophisticated conformational space sampling procedure which runs on a large parallel computer. The loop conformation generator searches through all of the conformational space and uses filters to eliminate unfavourable conformations. After all the modifications were carried out the entire protein was run through the energy minimiser. First polar hydrogens, then all hydrogens, and finally the water molecules from the bovine trypsin crystal structure were added to the model. Energy minimisation continued until the first derivatives from the Newton-Ralphson algorithm had become negligibly small.

Item Type: Thesis (MSc(R))
Qualification Level: Masters
Additional Information: Adviser: D N J White
Keywords: Biochemistry
Date of Award: 1997
Depositing User: Enlighten Team
Unique ID: glathesis:1997-75405
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 20:13
Last Modified: 19 Nov 2019 20:13
URI: https://theses.gla.ac.uk/id/eprint/75405

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