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Can the CCP4BB provide something like a website to upload pictures and then have the BB-ers just post the link in their email. Please! These attachments are clogging my inbox... Thanks much. Raji ---------Included Message---------- >Date: 31-jan-2008 03:58:44 -0500 >From: "Frank von Delft" <[log in to unmask]> >To: <[log in to unmask]> >Subject: [ccp4bb] Web site GOOD, Attachment BAD (was: salt sensitive complex) > >Actually, it's not book keeping, it's simple courtesy -- and not only on >a BB: an attachment is lazy, and a large attachment is downright rude. > >I am routinely stuck with a slow connection (travelling), and others are >*permanently* stuck with one. So please be nice... ;) > >phx. > > > >Anastassis Perrakis wrote: >> Dear all - >> >> Sorry to intervene on a 'book keeping' issue, but indeed over the last >> few months an increasing number of people (Jerry is not the first, so >> Jerry please do not take it personally) attach pictures etc. I think >> in a bb standard practice dictates to only use text - if illustrations >> are needed to explain the problem, you can put them in eg a web site. >> >> Some text like that was in the 'code of conduct' off ccp4bb in the >> past, but I could no longer find it. >> >> Thus apologies if I am wrong and policies have changed, but maybe the >> ccp4 crowd could tell us what is the suggested policy. >> >> And, if you really want to send an image please do bother to make it >> small. The initial posting had a 630k image, which it took me 1 min to >> make 20k and it still makes the point (attached so I can also violate >> the rules i am suggesting - I love inconsistency). >> >> Thanks, Tassos >> >> >> >> >> On Jan 30, 2008, at 20:11, Jerry McCully wrote: >> >>> >>> Dear All: >>> >>> Thanks a lot for the prompt reply on this topic of salt >>> sensitive complex. >>> >>> Attached please find one ITC final figure done under 25mM >>> Tris(pH8.0), 60mM NaCl. >>> >>> As mentioned before, the ionization of Tris will interfere with >>> the ITC experiments. >>> >>> Therefore I am sure of my binding results. >>> >>> Can anyone give me some comments on this ITC experiment? >>> Basically do these two proteins bind to each other? If so, how should >>> I improve the ITC experiments to get a similar affinity shown by >>> BIAcore(about 0.5uM)? >>> >>> Thanks again. >>> >>> Jerry >>> >>> ------------------------------------------------------------------------ >>> >>> Hi Jerry, >>> >>> Tris can cause problems, you are better off using something like >>> HEPES, and HEPES should be ok at pH 8. (Buffers with an >>> ethane-sulphonic acid group tend to be the best - those ending in >>> 'ES', so MES, TES and HEPES) >>> >>> FYI, the error on your K is bigger than the actual measurement - >>> 1.49x10^5 ± 1.5x10^5. >>> Signal to noise to is probably your enemy, which is making the curve >>> fitting difficult. Changing buffer may help this - there may be some >>> non-specific component to what you're seeing - increasing salt a bit >>> or dropping in something like 5% glycerol may help with this. >>> >>> Would you be able to post a jpeg/pdf of the curve? >>> >>> Regards, >>> >>> David >>> >>> >>> On 25/01/2008, Jerry McCully wrote: >>> > >>> > Dear All: >>> > >>> > Firstly I would like to thank many folks here for giving me great >>> > ideas several days ago. >>> > >>> > The following are some updates for this question. >>> > >>> > I did ITC experiments again using 25mMTris(pH8), 60mM NaCl(low salt >>> > condition). >>> > >>> > But things still turn out to be a little weird. >>> > >>> > I increased the concentration of both proteins(60uM in the cell and >>> > 1200uM in the syringe). At the end of the ITC, I saw a little of >>> > precipitation of both the proteins. >>> > >>> > Fortunately I can roughly fit the curve this time. However, the heat was >>> > still low, around 1Kcal/mole of per injectant. I am not sure about the >>> > fitting statistics. >>> > >>> > >>> > >>> > N 1.10 ±0.17 >>> > >>> > K 1.49E5 ±1.5E5 >>> > >>> > DH -893.5 ±213 >>> > >>> > DS 20.7 >>> > >>> > Was the enthalpy was offset by the ionization of Tris buffer? >>> > >>> > Can I use Hepes buffer around pH8 to do ITC? >>> > >>> > >>> > Welcome any comments about the statistics and suggestions on how to >>> > improve the ITC experiments.have a nice weekend. >>> > >>> > Jerry >>> > >>> >>> >>> >>> ------------------------------------------------------------------------ >>> Helping your favorite cause is as easy as instant messaging. You >>> IM, we give. Learn more. >>> <http://im.live.com/Messenger/IM/Home/?source=text_hotmail_join> >>> >>> >>> ------------------------------------------------------------------------ >>> Shed those extra pounds with MSN and The Biggest Loser! Learn more. >>> <http://biggestloser.msn.com/> >>> <test-ITC-012608.JPG> >> > > ---------End of Included Message----------