buried regions may become reexposed and overestimated;
(2) apolar stretches that were protected by nearby structural
gatekeepers could be overestimated; (3) vanishing of a key
apolar amino acid that determines an aggregation prone region
would cause underestimation. As a conclusion, the probability
to predict nondesired artifacts increases as the number of
sequence gaps increases. It is the user responsibility to evaluate
if the missing region of the protein structure is crucial for the
prediction of protein aggregation and if, accordingly, it can lead
to wrong A3D score values.- If the project has no associated name, A3D will automatically
label them using an internal standard code. When performing
predictions with large data sets or a large number of mutations
it is highly recommended to name the different projects
individually. - When deciding between the static and dynamic modes of A3D
the user should gather as much information as possible on the
target protein to balance the benefits and disadvantages that
might result from running the protein in any of the two modes. - Amino acids without 3D coordinates in the PDB file are not
taken into account in predictions and are not visible in the
mutation tool of A3D. - A3D tolerates as many mutations as the user considers appro-
priate to scrutinize. However, it has to be taken into account
that an excess of mutations might promote the attainment of
nonnative conformations and this effect is not considered by
the algorithm in its present form. - It is important to save the URL or bookmark the project
window if we have chosen the option “do not show my job on
the results page.” Even though the introduction of a user e-mail
address is not mandatory, it is highly recommended for those
projects that are not visible in the queue list. When the run
finishes, the system sends a link that redirects the user to the
specific job page, where the results are available. Skipping this
step might result in the user not being able to access the
prediction results. - “Total score value,” absolute value resulting of the sum of all
A3Dscore;“Average score,” results from dividing the total score
value for the number of total amino acids in the protein. It also
takes into account those buried residues with A3Dscore¼0;
“Minimal score value,” amino acid with higher predicted solu-
bility within the structure and “Maximal score value,” amino
acid with higher aggregation propensity within the structure. - Usage of the score-table to mutate residues is recommended
when comparing a point mutation to thewild typeversion of
the protein, since the energetic correction performed by FoldX
440 Jordi Pujols et al.