Arabian Came alpha-1-microglobulin/bikunin precursor (AMBP), mRNA

[SULTAN-ALHARBI]

• please download and use the sequence (alpha-1-microglobulin/bikunin precursor (AMBP), mRNA): AMBP_protein.txt
• First find Open Reading Frame (ORF) then translate the AMBP mRNA to Protein then do the following:
  • (1) multiple sequence alignment and analysis of phylogenetic relationships
  • (2) prediction of the secondary and 3D structure of protein
  • (3) physical and chemical properties of protein
• you will receive an invitation form ShareLatex to be one of the work collaborators to edit the draft paper of Arabian Camel AMBP protein: link to Arabian Camel AMBP draft: Arabian Camel AMBP draft
• please don't hesitate to post a question or an enquiry, the team members will be happy to help
• please respect the following Deadlines:
  • [x] perform & generate results of in silico analysis only: 24 August 2017
  • [x] write section of results of in silico analysis only: 17 September 2017
  • [ ] write section of methods of in silico analysis only: 1 October 2017
  • [ ] write section of introduction: 12 October 2017

All the best

[MManee]

For more information about how you can perform and generate your results, please check this issue #14

[1sar2003]

Dear all
I used ncbi ORF finder to determined AMBP ORF with Search Parameters below, and I end up having 14 ORFs, should I change minimal length or codon to have less result or keep it stander as it is ?

[SULTAN-ALHARBI]

Hi @1sar2003
Open Reading Frame (ORF) is the region in your mRNA which will be translated to protein, almost all proteins are started with Met(methionine ) amino acid this amino acid has ATG(AUG) codon. so we should use "ORF start codon to use : "ATG" only this is first. second: if you search for AMBP protein length for any well-annotated species, you will obtain that the length of AMBP protein should be around 350 amino acids. For example Human AMBP protein length is 352 amino acids and Caw AMBP protein is 334 amino acids. Our Arabian Camel AMBP protein length will be around those lengths. You should end up with only one ORF

[MManee]

• it is recommended that you use geneious software
  • download Link: http://www.geneious.com/
  • licensee: SULTAN ALHARBI
  • license Key: 1BFA-15C8-99BC-1DF9-60C1
  • valid from 07 July 2017 to 07 July 2020
  • you could use trial version first
• @malsaeed please let me know if you can help @1sar2003 in how to use geneious software for the same analyses

[malsaeed]

@MManee @1sar2003

We already have small call about the work today

And I'm glad to help any person with geneious

We will see if we can meet tomorrow

Regards

[1sar2003]

thank you all for the respond the license has been activated the maximum number of times so I will use the trial version
@MManee @malsaeed @SULTAN-ALHARBI

[SULTAN-ALHARBI]

That is weird :( Because we have 5 license users and we just used one so far. I'll check with Geneious team what the problem.

[1sar2003]

Hello
I used geneious to perform my task, the length of Arabian Camel AMBP protein is 352 aa end up with 1 ORF(length 1059) and I followed the next :

1/Translate AMBP mRNA to Protein (BLAST):

2/multiple sequence alignment :

3/ phylogenetic relationships (Tree)

I will continue to update the result soon, just need to double check

[SULTAN-ALHARBI]

Great to see a quick movement let's take your work one by one 1/Translate AMBP mRNA to Protein: yes, Arabian Camel AMBP protein is 352 aa. 2/multiple sequence alignment : you can do this task in two ways A) using Arabian Camel AMBP mRNA (1059 bp) and this is what you already did. Here you are comparing Arabian Camel AMBP mRNA with mRNA from other species. B) using Arabian Camel AMBP protein 352 aa and comparing it with AMBP proteins from other species. and this what we actually want all published articles use this method. so make a multiple protein sequence alignment. 3/ phylogenetic relationships (Tree): it is a good tree, we can see our Arabian Camel grouped with Bactrian camel (Camelus bactrianus) and alpaca (Vicugna pacos). but we need to construct our phylogenetic tree using protein sequence alignment

the the best

[1sar2003]

Dear @SULTAN-ALHARBI how should I obtain the Arabian Camel AMBP protein seq alignment in geneious? is it by select ORF and blast it in the protein data bank?

[1sar2003]

like when I blast the trans protein under the Nucleotide collection I end up have a list of organisms does not include my dromedaries Camelus, and the same thing applied in most of Blast database

Tree

By the way, I obtained the translated protein by click Sequence / Translate

[SULTAN-ALHARBI]

Hi @1sar2003 First/ Select the region in the sequence that you want to translate Second/ go to icon called sequence ---> translate . You will get your translated sequence as separate file. you this seq in multiple alignment then phylogenetic tree

[1sar2003]

@SULTAN-ALHARBI this is exactly what I did before, the only thing that I change now is running the blast program with (tblastn) instant of (blastp) and I get what I need

1/Multiple protein sequences alignment:

2/phylogenetic relationships (Tree):

3/prediction of the secondary AMBP protein structure: 8ba26956-8328-11e7-9477-00163e110593.AMBP.psipred.pdf

4/prediction of the 3D AMBP protein structure:

[1sar2003]

5/ Isoelectric point for (AMBP):

6/ physical&chemical properties of (AMBP) by using Kolaskar & Tongaonkar method only:

7/Prediction ordered and disordered regions of (AMBP) protein : Prediction ordered and disordered regions (AMBP) alpha-1-microglobulin:bikunin precursor.pdf

8/Conserved domains in (AMBP) protien seq:

Have a nice weekend.

[MManee]

• @1sar2003, this was an incredible achievement accomplished in a short period of time
• so we will have to move forward to the next step, which is write up
  • more reference papers were added here: gene in silico analysis
• we will help writing the manuscript:
  • [x] results - @1sar2003, @AbuMohammad and @SULTAN-ALHARBI
  • [ ] methods - @1sar2003, @AbuMohammad and @SULTAN-ALHARBI
  • [ ] introduction - @SULTAN-ALHARBI
  • Deadline: 12 October 2017

[1sar2003]

Dear all @MManee @SULTAN-ALHARBI
I'm trying to reconstruct multiple alignments and phylogenetic tree and I facing some of the species didn't start with M !! is anyone can explain why? and most importantly how to resolve it? *Most of the feature on geneious is restricted

[SULTAN-ALHARBI]

Dear @1sar2003
Great what you did, it seems to me that you do not have any problem with making multiple alignment and constructing a phylogenetic tree. If you find any species that doesn't fit with your exceptions just remove it. ( see the one in LaTex )
I think you should move in to 2nd and 3D structure and domains of AMBP protein . we should write and read more about AMBP structure

• About Geneious software we just purchased new license for you, I will send it to you when I got it from Geneious team. All the best

[SULTAN-ALHARBI]

Dear @1sar2003 @malsaeed @MManee This is the new License Key for Genious

Licensee: SULTAN ALHARBI License Key: 1BDA-15EE-1B52-92E1-C6A1 Valid from 03 October 2017 to 03 October 2018

[1sar2003]

Dear @SULTAN-ALHARBI thank you for your comment, I think I already write about the domains region and regard 3D structure I admit I having a difficulty of understanding the basic of it and will continue trying my best with it.

best regard

[SULTAN-ALHARBI]

Dear @1sar2003 The information of domins'A1M is very good I will work on it these days. we need one more step to confirm them as 3D structures. I know, it's easy to annotate novel 3D. we will be side to side helping each other

[1sar2003]

Dear @MManee @SULTAN-ALHARBI In the writing process, I have some thoughts I would like your consultant in it, should I mention the biosynthesis of the two proteins in the cell and the path they follow to express.Second which category the protein inhibition of the protienes falls? is it in discussion or introduction section?

[SULTAN-ALHARBI]

@1sar2003 Hi Sara As most literatures mention those two Protein (A1M and Bikunin are co-synthesis = coming from same mRNA) our scope in this paper is to annotate the structural properties of those proteins as well as their biophysical features. we can briefly mention their biosynthesis pathways just to support any structural or biophysical characterizations. For example\ the section where we want to explain the trypsin interaction site or glycosylation site on the protein. about where to explain such information either on introduction or discussion, it depends on the coordination of the paper.

All the Best

[MManee]

• @1sar2003, we need to put a plan to fill the current gaps of this work
• we may not consider sequencing AMBP, and use the predicted sequence