Parent-map: analysis of parental contributions to evolved or engineered protein or DNA sequences

Full text

Parent-map: analysis of parental contributions to evolved
or engineered protein or DNA sequences
Damien Marsic1
1Porton Biologics, 388 Xinping Street, Suzhou Industrial Park, Jiangsu 215021, China
DOI: 10.21105/joss.02864
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Submitted: 18 November 2020
Published: 23 January 2021
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Summary
Parent-map analyzes protein or DNA sequences which are derived from one or multiple parent
sequences, and shows parental contributions as well as dierences from relevant parents. Orig-
inally developed to analyze capsid protein sequences obtained by directed evolution, parent-
map can be used in any case where variant sequences are to be compared to parent sequences
from which they are derived. Parent-map detects sequence shuing as well as substitutions,
insertions and deletions, and displays results in user-friendly formats. Parent-map is an open-
source, platform-independent Python 3 script, available as a Bioconda package as well as a
Windows program.
Source code: https://github.com/damienmarsic/Parent-map
Python package: https://pypi.org/project/parent-map/
Bioconda recipe and package: http://bioconda.github.io/recipes/parent-map/README.html
Windows installer: https://sourceforge.net/projects/parent-map/
Documentation: https://parent-map.readthedocs.io/
Statement of need
Adeno-associated virus (AAV) capsid directed evolution projects typically generate multiple
enriched variant sequences after 2 to 5 rounds of selection starting from complex capsid
libraries. For libraries developed from a single parental serotype, through random peptide
insertion at a specic position or surface loop diversication in well-dened variable regions
for example, a single multiple alignment of all enriched variant sequences against the parent
sequence conveniently shows how each variant diers from the parent. However, when more
than one parental sequence is involved, such as when dierent libraries are mixed together, or
when a library design involves DNA shuing from several parents, such alignments can quickly
become illegible, particularly when the complete capsid gene is sequenced. In such cases, in
the absence of appropriate software tools, each variant needs to be separately aligned against
all possible parents, a time-consuming and cumbersome process. An added diculty in the
case of shued libraries is that, because of high sequence homology between parents, multiple
regions will share sequence identities with more than one parent, complicating attempts at
comprehensively dening the variant sequences in terms of parental contributions. To date,
SALANTO (Herrmann et al., 2019) seems to be the only relevant publicly available software.
However, it only applies to shued libraries, and its user-friendliness is limited as it requires
the user to perform a multiple sequence alignment beforehand, and to further process the
data manually after analysis. The software described in this article, parent-map, provides a
user-friendly and comprehensive solution. It can be used with sequences derived from any
Marsic, D., (2021). Parent-map: analysis of parental contributions to evolved or engineered protein or DNA sequences. Journal of Open Source
Software, 6(57), 2864. https://doi.org/10.21105/joss.02864
1

type of library, or even with naturally-occurring mutants or rationally engineered variants. It
is not limited to protein sequences. It only requires one le containing the variant sequences
to be analyzed, and one le containing parental sequences, without any prior manipulation. It
generates a set of ve les covering most end-users’ needs, in directly usable formats. Finally,
although it was developed to address a need in the eld of AAV capsid directed evolution,
parent-map can be used whenever protein or DNA sequences, whether originating from natural
evolution, directed evolution or rational design, are to be compared with one or more possible
parental sequences.
Methods
Parent-map was written under Python 3.7 as both a command-line interface (CLI) and a
graphical user interface (GUI) application, by allowing parser modules argparse and Gooey to
coexist within a single le (the GUI will start if no argument is present, while any argument
will cause parent-map to start in CLI mode). A parent-map Python package was created and
uploaded to the Python Package Index (PyPI) according to packaging instructions. A parent-
map Bioconda (Grüning et al., 2018) recipe based on the PyPI package was written and
submitted according to instructions. A stand-alone Windows executable and its installation
program were created using respectively PyInstaller and Inno Setup. The documentation was
written using Sphinx.
Implementation
Parent-map is a platform-independent Python script that generates a set of ve output les
from two input les. Input le names and options can be entered as arguments at launch time,
resulting in parent-map running in CLI mode, or within the GUI, which starts if parent-map
is launched without arguments. This exibility allows parent-map to be deployed in a variety
of settings, as a simple desktop application or even as a bioinformatics pipeline component.
The rst input le contains the variant sequences, typically the most frequent or the most
enriched sequences obtained at the completion of a directed evolution experiment. The other
input le is a set of potential parental sequences to the variant sequences. The most useful
les generated by parent-map, particularly in the case of variants derived from DNA shuing,
are parental contribution maps (le names ending in –par.txt and –par.html, the latter being
a colorized version of the former). Instead of all possible combinations, the simplest map
that can accurately describe the variant is shown, using as few parents and as few fragments
as possible. Other output les include a statistics le summarizing the variant sequences
main features, a sequence denition le comprehensively dening each variant in terms of its
parents, and an alignment le showing how variants dier from their common parent.
Parent-map can be tested using the provided variant and parent sample les, based on available
literature describing evolved and rationally designed AAV capsid variants. Variants AAV-DJ
(Grimm et al., 2008), AAV2.5T (Excoon et al., 2009), NP84 (Paulk et al., 2018) and
OLIG001 (Powell et al., 2016) are derived from shued DNA libraries. Variants AAV-F
(Hanlon et al., 2019), AAV-PHP.B (Deverman et al., 2016), 7m8 (Dalkara et al., 2013)
and rAAV2-retro (Tervo et al., 2016) are derived from peptide insertion libraries. Variants
SCH2, SCH9 (Ojala et al., 2018), LI-A and LI-C (Marsic et al., 2014) are derived from more
complex rationally designed libraries. Variants AAV2i8 (Asokan et al., 2010) and AAV2-sept-
Y-F (Petrs-Silva et al., 2011) were rationally designed. Using default settings, parent-map
correctly identies single parental contributions from AAV9 for variants AAV-F and AAV-
PHP.B, single parental contributions from AAV2 for variants 7m8, rAAV2-retro, LI-A, LI-C,
AAV2-sept-Y-F, and multiple parental contributions from AAV2, AAV8 and AAV9 for AAV-
DJ, from AAV2 and AAV5 for AAV2.5T, from AAV2, AAV3B and AAV6 for NP84, from
Marsic, D., (2021). Parent-map: analysis of parental contributions to evolved or engineered protein or DNA sequences. Journal of Open Source
Software, 6(57), 2864. https://doi.org/10.21105/joss.02864
2

AAV2, AAV6, AAV8 and AAV9 for OLIG001, SCH2 and SCH9, and from AAV2 and AAV8
for AAV2i8. Parent-map also correctly detects peptide insertions FVVGQSY for AAV-F and
TLAVPFK for AAV-PHP.B, both at position 588, and peptide insertions LALGETTRPA for
7m8 and LADQDYTKTA for rAAV2-retro, both at position 587. Finally, parent-map correctly
identies substitutions A to T at position 457 for AAV-DJ and at position 582 for AAV2.5T,
substitutions K to E at 532 and R to G at 585 for NP84, E to K substitution at 532 and
unmatched H at 726 for OLIG001, substitutions I to T at 240 and V to I at 718 for 7m8,
substitutions N to D at 382 and V to I at 718 for rAAV2-retro, the 14 and 4 substitutions for
LI-A and LI-C respectively, as well as the 7 Y to F substitutions at 252, 272, 444, 500, 700,
704 and 730 for AAV2-sept-Y-F.
A comprehensive description of parent-map is provided in the documentation.
Acknowledgements
We thank Yan Chen and Oleksandr Kondratov for testing parent-map and providing valuable
feedback.
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Marsic, D., (2021). Parent-map: analysis of parental contributions to evolved or engineered protein or DNA sequences. Journal of Open Source
Software, 6(57), 2864. https://doi.org/10.21105/joss.02864
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