Well, look at that! I had missed that one. Thank you Dan.
Ahh, those Ig folds. They are like dougnuts. Is there anything they
can't do?
Interesting that Tan et al. arrived at their conclusions from a very
different line of evidence. They seem to have missed the covariance
with the silent site pointed out by Tang et al.?
1xak is from original SARS, and I must say not great statistics despite
being at 1.8A. Still, it is a start.
From the literature cited by Tan et al. it appears ORF8 has been
suspected as pathogenic for some time, but for widely different
reasons. One paper says it is a transcription factor that up-regulates
chaperones, another says it is a secreted imunosuppressor.
So, my question remains: why haven't we solved it yet?
-James Holton
MAD Scientist
On 3/21/2020 9:12 AM, Rigden, Dan wrote:
Hi James
5o32I is not a homolog of ORF8 - the BLAST e-value is insignificant.
In fact, rather than the EGF-like fold of 5o32I, ORF8 has an Ig-like
fold similar to ORF7 (for which there is a structure; 1xak).
https://toolkit.tuebingen.mpg.de/jobs/2717885_1
I must say I got quite excited seeing that until I noticed this
pre-print which tells the whole story very nicely, including that key
position 84....
https://www.biorxiv.org/content/10.1101/2020.03.04.977736v1
Best wishes
Dan
------------------------------------------------------------------------
*From:* CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of
Patrick Shaw Stewart <patr...@douglas.co.uk>
*Sent:* 21 March 2020 15:41:17
*To:* CCP4BB@JISCMAIL.AC.UK
*Subject:* Re: [ccp4bb] CCP4BB vs COVID19
James, this isn't conventional structural biology, but may be of
interest, and I haven't been able get any mainstream virologists to
think about it.
The protein sequences are obviously of interest, but so are the RNA
sequences at both ends of the Covid genome, which have conserved
secondary structure. A few years ago a paper came out suggesting that
wild-type influenza has multiple "RNA thermometers", which may play an
important role in the tropism of influenza. Similar mechanisms may
exist in other respiratory viruses, including Covid.
My take on this, and the relevant papers, are below.
Good luck to everyone and stay well,
Patrick
https://oldwivesandvirologists.blog/Covid-19-and-the-trade-off-model-of-selection/
My paper in /Medical Hypotheses
/http://douglas.co.uk/f_ftp1/ShawStewart_final_1-s2.pdf
Narberhaus, Franz, Torsten Waldminghaus, and Saheli Chowdhury.
"RNA thermometers." /FEMS microbiology reviews/ 30.1 (2006): 3-16.
Chursov, Andrey, et al. "Specific temperature-induced
perturbations of secondary mRNA structures are associated with the
cold-adapted temperature-sensitive phenotype of influenza A
virus." /RNA biology/ 9.10 (2012): 1266-1274.
Yang, Dong, and Julian L. Leibowitz. "The structure and functions
of coronavirus genomic 3′ and 5′ ends." /Virus research/ 206
(2015): 120-133.
On Fri, Mar 20, 2020 at 10:59 PM James Holton <jmhol...@lbl.gov
<mailto:jmhol...@lbl.gov>> wrote:
You might think that as a structural biologist you won't be able
to do
much about COVID-19 anytime soon, but that is not true. Yes,
real-world
therapeutics and vaccines take time, but we have already seen just
how
fast we can get started. There are 21 PDBs already and some even
have
bound ligands. Good job Frank et al. BTW! And my personal thanks to
all of you out there who are already hard at work on this.
I believe this forum is an ideal place to share information and
ideas on
the structural biology of SARS-CoV-2 as we move forward. It's a big
virus, but there are not that many proteins in it. If all of us
independently do the same bioinformatics and literature searches
and end
up trying exactly the same thing in every lab all over the world,
then
that would be more than unfortunate. To that end, I am personally
interested on ORF8 for reasons I will go into below. Has anyone
tried
to solve it yet? What happened? Didn't express? Bad diffraction?
What? Do tell.
Some of us, as you may have heard, are stuck at home, our
beamlines and
labs dark while we shelter-in-place. That doesn't mean our hands are
tied. We are still allowed to think. The fraction of the human race
that has a snowball's chance in Hades of figuring out this bug is
very
very small. Structure may be your main skill set, but you are
still a
biologist. Do you know how to run a PCR machine? Do you know how to
pipette? You might think that anybody can do it, but that is
really not
the case. Ever trained a new student on sterile technique? How many
days did that take? Now remember that your student was no dummy and
already studying biology. Everyone reading this will make an
excellent
volenteer at the very least. I'm not saying this to belittle the
average human, only to say that we scientists, moving in the
circles we
do, often forget that we have uncommon capabilities.
For example, I also believe we can be useful in assay development.
The
void left by the dearth and delay of test results has been filled
with
fear, and that is a big problem. The tests, as defined, are
straightforward, but also extremely regimented like any good
laboratory
protocol should be. The US CDC's instructions for academic labs
are here:
https://www.cdc.gov/coronavirus/2019-nCoV/lab/index.html
My question is: how can this test be made faster, using more
commonplace
supplies, in high-throughput mode and still valid? Not just for
clinical but for academic use? I think more than a few people on
this
list could be regarded as experts in making a complex biochemical
task
faster, more efficient, high-throughput and nonetheless valid. Yes,
there are other people who do virus testing for a living, but
right now
they are all rather busy. Maybe if we put our minds to it we can
help?
As for why ORF8. I am basing my interest on the bioinformatics
done in
this article: https://dx.doi.org/10.1093/nsr/nwaa036
<https://dx.doi.org/10.1093/nsr/nwaa036>. Search for
"T8517C" and you will find what I'm talking about. The authors found
two "types" of SARS-CoV-2. They call them "S" and "L" because the
only
conserved amino acid change involved is S84L in ORF8. The "S"
type is
believed to be the ancestor of "L". What is interesting is how
tightly
linked this mutation is to a silent mutation on the other end of the
genome: the "L" type has a faster codon for Ser in ORF1. Such tight
coupling (r^2=0.945) means there must be significant selective
pressure
preventing both of these mutations occurring in the same virus at the
same time. That, I believe, is interesting. Espeically since
they are
so far apart I expect this selective pressure might work in trans:
as in
a super-infection. That is, the S and L genome types may interfere
with
each other.
The authors fall short of claiming evidence of interference upon
super-infection, and indeed they have already been criticised for
calling "L" the "aggressive" type. But it is still interesting and
points a finger at ORF8.
ORF8 has only one homolog in the PDB: 5o32 with 25% identity over a
stretch of 60 residues. This homologous region contains the S84L
site
(Val I544 in 5o32). I had a quick look and appears to be a
cavity-filling mutation to me. Not very big, but maybe something
could
fit in there. To be sure we'd need a structure of ORF8.
Good luck to you all, and stay healthy.
-James Holton
MAD Scientist
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