Calculating Isoelectric Point of Proteins (Example)

Donate here: www.aklectures....
Website video link: www.aklectures....
Facebook link: / aklectures
Website link: www.aklectures.com

[@aarondesalvio]

Thank you so much, this was explained incredibly well! One piece of advice for anyone who's confused: keep in mind whether or not the side chain is neutral or positive in the protonated form. This is relevant for the COO- group, where it is NEUTRAL when protonated, as opposed to an NH3 group that is POSITIVE when protonated.

[@SouthpawC]

thanks!

[@JuanRomero-xe2lf]

KZbin Channels like this one-- (among a few others) are the future of learning. Thank you.

[@lazer1235]

Very well explained, best video I have come across on youtube that explains this principle thus far

[@AKLECTURES]

lazer1235 Thanks! I appreciate that :)

[@elenakhachaturyan8146]

+lazer1235 Totally agree!!!

[@charmainemoya2524]

Wow, you're a LIFE SAVER. While I spent hours on homework trying to read my book and notes and still scratching my head, you make everything clear in less than 15 minutes per lesson. Love all your videos!

[@absenzz]

Not all heroes wear capes...

[@AKLECTURES]

+Fab Gon Thanks bud! :)

[@superoxidedismutase5757]

some wear AK lectures t-shirts

[@thubelihlesithole5184]

I'm a bit confused, I'm not quite sure why you concluded the two acidic amino acids as neutral and not positive.

[@official.rajarshidutta]

exactly my doubt

[@kritankunwar4776]

@@official.rajarshidutta I think its because glutamate and aspartate was negatively charged hence oxygen has a negative charge to start with so when you add hydrogen that makes it neutral.

[@pfeliciano4062]

So if the PH is within 1 unit away from the Pka the charge remains neutral. You see that we chose 3.5 as a guess for the PH, and the difference between that Ph=3.5 and Pka-4.1 is 0.6 and this is within the range of 1 unit away. You cannot have greater or less than that 1 unit. also if you are wondering why we did not just use a PH of 4.1, that is because for our second step we need to pick the pka values that are above and below our PH estimate, so as we knew we needed to pick a value within 1PH unit away from that Pka, we chose the PH close close enough to 4.1. Hope this helps and God bless your studying!

[@sonjak8265]

@@pfeliciano4062 thank you

[@yazan4mjad]

Simply when pH goes lower than pKa of the carboxylic group which is 4.1, the carboxylic (-COO-) group will be protonated (-COOH) and it will have no charge, thus it is considered neutral.

[@reubent5780]

AK LECTURES Hey great video ..one thing that i'm a bit stuck on is why pka of 4.1 was neutral at pH 3.5 for the first example and why it was neutral for pka of 8.3 at ph7 in the 2nd example. thanks

[@marchizaaa]

reuben Thomas george At pH=3.5, the Carboxylic group (pKa 4.1 in the picture) is neutral because is protonated,so you have a COOH group without charge. At pH=7 is the same...the SH group of the cysteine (pKa 8.3 in the picture) is neutral because it can do only 2 bonds, like an hydroxyl group ( OH), and because of this is neutral (it's difficult that O and S do 3 bonds,usually on organic molecules they are unstable in this state)...it has no charge. If we were at pH= 10 , the cysteine would be S- with a negative charge ( the -SH group would lose a proton because of OH anion forming H2O). I hope it is clear now. See ya!

[@thepowerofpositivity8075]

+reuben Thomas george Jus follow the simple rule if (1) the compound is a weak acid then the protonated form will be uncharged (AH) and the deprotonated form (A) will be negatively charged;(2) the compound is a weak base then the protonated form (BH) will be positively charged and the deprotonated form (B) will be uncharged. The tendency a compound will donate or accept proton depends on pH of the solution. Hope this help!

[@the.angeltaylor]

@@marchizaaa I don't really see where the COOH group is pronated

[@marchizaaa]

@@the.angeltaylor a COO- group (pKa = 4.1) gets protonation at pH 3.5. Therefore becomes COOH group, a neutral charged one.

[@the.angeltaylor]

@@marchizaaa thankyou so much!!

[@thepowerofpositivity8075]

Jus follow the simple rule if (1) the compound is a weak acid then the protonated form will be uncharged (AH) and the deprotonated form (A) will be negatively charged;(2) the compound is a weak base then the protonated form (BH) will be positively charged and the deprotonated form (B) will be uncharged. The tendency a compound will donate or accept proton depends on pH of the solution. Hope this help!

[@andrewcho9779]

Thank you so much! I got very confused on this section and my professor wasn't of much help on figuring out how to to this but looking at this, I realize that it is much more simple than we think!

[@chillstep4life]

Amazing! Very easy to understand, very well organized! Cant say enough about how clear and concise your lecture is. Thanks for the video.

[@Horsin4years]

This was super helpful. I like that you explain the steps very clearly and make a point to separate them out.

[@cambokid16]

At first your videos were intimidating at first glance by the excessiveness, but omg it's very simple, straight to the point, and the explanation is beautiful, thank you!

[@pj10red]

Thank you soo much! My first Biochem exam is tomorrow and you saved me!!

[@TonilaineC]

After watching this, I might just pass my biochem exam. Thank you so much 😭😊

[@joseibarra9808]

Your videos are so helpful. Great for the new MCAT.

[@tylerlynn2655]

this guy is the only reason I made it to pharmacy school

[@funnysjoker]

Wow great lecture! Thanks so much! Keep the videos coming! I watched this on the bus, and it was very easy to follow. Thank you!

[@zay7968]

Thank you so much! I have my final exam tomorrow and this helped a lot! Hope u succeed in whatever you do.

[@AKLECTURES]

+Shadow walker Thanks! I appreciate that! best of luck on your exam, let me know how it all turns out!

[@britforlife32]

I didn't understand how to calculate pI before this, but now I do! Thank you so much, it's much appreciated

[@catherinepaciotti3800]

Hi, thanks for the video. Did you forget to take into account the pka of cysteine?

[@Nanyaq]

Perfect! Thank you so much --this really helped me before my biochem exam! :)

[@penelopetaylor8384]

You are a true hero. THANK YOU for these amazing lectures.

[@YaacovPsuturi]

I love u man, your explanation is perfect!!!

[@TheToxicMegacolon]

AK saving the day again for the 6th semester lol!

[@jalehnikfarjam9844]

Thank you for the clear explanation!

[@jacksonwestaway3376]

Your videos are fantastic! Rlly appreciate them and you; getting me through MBLG =)

[@amerain1729]

This is very helpful. Thank you!

[@pratibhamali3845]

this is very helpful to me.thank you so much.

[@abayady1]

This is a great video, thank you! However, I have a question, why don't you count the charges of amino acids individually? For example, Aspartate and Glutamate are both negatively charged aa, but you do not address their charge, and start as if they were neutral? My professor does count them. Would love to hear your feedback

[@blehm8400]

Basically, the easiest way I found to think about it is this. At a very low pH (e.g. pH 1) there is a high concentration of H+ ions (this is what pH measures). Therefore, anything that can accept a H+ ion will do so. This includes aspartate and glutamate which you'd normally think of as being negatively charged due to the COO- group they both have at the end of their side chains. When we raise the pH to 4.1 (the pKa value of aspartate and glutamate) they will accept a H+ ion and the COO- group will become COOH. So basically, start from a low pH , where everything that can be protonated (contains a H+ ion) is protonated. Look at the overall charge of the peptide. Then increase the pH and each group with a pKa value will be deprotonated (lose a H+ ion) at its pKa value. At each pKa value, note the new overall charge of the peptide. Keep doing this until you find the 2 pKa values that you will average to get the pI (isoelecric point). Hope that clairifes it.

[@crystalguerreromorris7102]

Your explanation was flawless, I finally comprehend!!!

[@shaboatrad4066]

Aw.. why not using the pka of S-H which was 8.3 instead of 8.0 ?! Isn't that wrong?

[@gorkemnailaydn1524]

Dude you are awesome. Thank you so much for this understandable and good lesson.

[@asie5651]

this guy had me thinking he's left handed for one minute 29 seconds.

[@MultiSamSami]

Man... I was looking for this. Thank you so much.

[@AP-lj5qe]

amazing video, very clear explanation. I am glad to watch this video because my professor explanation is sucked.

[@omarrosario1489]

Sir, why I did not find you earlier? Excellent video, may God protect you wherever you are and gives you health and happiness. Good vibes.

[@TheMachomaniac]

Dont guess. Start with a fully protonated protein and solve towards a higher pH until you find the answer, a neutral peptide. The you find the pKa range and can solve the pI.

[@crackers0413]

Hi, I’m a little confused. The pka of the NH3 of Cysteine is 10.7, so averaging 4.07 of glutamic acid and 8.37 of cysteine gives 6.22

[@ahalyalenka7358]

wow......... Lheninger ate my head....take my Namaskaar...

[@simrankaur11]

That was very well explained! Thank you.

[@syyylvo]

Very clear and coincise!

[@jacobboeckelman3691]

This helped out so much. Thanks!

[@caiquebarreto6498]

Thank you for this awesome explaination!!!

[@enricotortelli3642]

You literally saved my life!

[@tomasaguilar3017]

You make this too easy to understand it makes me question if Biochemistry is even a difficult subject......

[@remyakin7198]

You just saved a life!! Thanks

[@shivansh709]

Thanks bro! Helped me in my aits!

[@kareemm8608]

My issue with a pH of 3 is that there will be a fractional charge in the COO- terminal so it won't necessarily be -1.

[@the.angeltaylor]

Why would the charges for the first example be neutral?

[@buluchka]

I feel this will save my biochem grade, Thank you.

[@salam495]

thank you alot !

[@walaahussein462]

Thanks for the video. How to calculate PI for modified peptides such as lantibiotics?

[@roblingbling]

great videos!. If you figure out that a 15kDa protein has a PI of 9.71, and then it forms a homodimer. Does that affect the PI?

[@AndreasChristodoulou99]

I love you for this video sir

[@chanchalmaji1456]

Again thanks for this

[@Kka-ong-the-cat]

Super helpful

[@DefenderX]

Is the pka value for each functional group in each amino acid accurate? Or do they vary depending on the peptide sequence and potential folding?

[@monday2471]

do you exclude the pKas of the terminal groups when calculating the pI?

[@RandomNooby]

@AK Re COVID Why is the death rate in Italy in 0-18 year olds less than 2% with everyone else roughly evenly divided into 3 equal groups. With many young adults to 50 year olds suffering from a cytokine storm, what are your thoughts?

[@SoraKimchi]

Thank u so much man❤

[@zena1212]

I finally get it thank you so much!

[@junczhang]

thanks!!!

[@lovelly2]

For problem 2, I don't understand why cysteine did not gain a positive charge. The pH is less than the pka which means it should have been protonated. I understand the charges for all the others but do not understand your explanation of cysteine.

[@the.angeltaylor]

This is confusing, why do the rules apply to one part of the protein and not the others? What makes the Cysteine side chain group different that the others? What makes the pka of 4.1 in the first problem different than the other parts of the protein? Can somebody break this down please?

[@lorcanokane5128]

Much better than my professor

[@yaarithanan1994]

Thank you very much!

[@mauimanman5498]

In the second problem where the pka is 8.3 why is it neutral?? wasn't it suppose to be positive because the pka was above the ph?

[@soumyamajumdar7929]

can you please find the pI for ala-arg-gly.

[@nunuchaa]

what kind of protein doesn't start with methionine

[@sharmaeaselearningbiologyc5277]

Sir thanku from india

[@dblkarno]

how do you know that the pKa of the H2N of the Asp. is 8.0?

[@lixx1414]

My teacher gives them to us

[@ansammkh1681]

thank you

[@husamidrees4609]

Great video.

[@chamandeepkaur2177]

Thank you so much. You're the best :-)

[@leonardmulenga6021]

I still don't understand when it comes to the estimations How those where becoming neutral instead of +

[@celestial4646]

thanks from a local ub student ;)

[@anyamiller514]

Why does Gly have a pk value? I wasn't aware that it had an ionizable group.

[@anyamiller514]

Shouldn't it just be ignored in the calculation?

[@jadzgburella2068]

Thanks ak lecture!!

[@jadzgburella2068]

Getting my yoni all steamed up

[@hasanciftci9432]

Amazing.

[@MsTommyknocker]

I tried calculating the isoelectric point of the polypeptide with the following sequence: KRHKKDE. I got the pI to 11.51, but expasy protein parameter calculator got it to 9.74 or something like that. Could you clarify???

[@jdevola999]

+MrTommyknocker I agree -- it seems like none of these calculators agree. Why is that?

[@czechmeoutbabe1997]

I don't understand why sometimes he says the higher pKa means that it has a *positive charge* at a pH, but sometimes it has a *neutral* charge at that same pH value. They're both above the pH, I don't see the difference.

[@gabrielwinces6606]

well explained

[@pathbasics]

So SH can't gain any more protons when its pka is above the ph? 12:35

[@niloofarfam4294]

Thank you!!!!!

[@_Maya_Andrea]

THANK YOU SO MUCH OH MY GOD

[@TheBollywoodCritic]

May god bless you with a thousand sons.

[@fukpoeslaw3613]

S.S .A Because we are not all feminists.

[@tatendawarima4607]

superb, thanks a lot!!!!!

[@AKLECTURES]

Tatenda Warima you're welcome :)

[@davehileri7072]

Great👍

[@joancalatayudvernich1237]

GREAT!

[@davehileri7072]

Ty

[@lixx1414]

QUESTION, SOMEONE HELP PLZ :D why are the pka=4.1 neutral with the guess of ph= 3.5 ?? shouldnt they both be positive...bc theyre above the guess??? leaving the over all charge to be +2?? help plz!

[@lixx1414]

+AKLECTURES

[@franciscoisidro3622]

I srsly don't know how you got the pKa of NH3 as 8

[@ElizabethRey97]

Thanks! I didn't even start college and I perfectly understood this :)

[@harinisuresh456]

How come you have solution for all my assignments!!

[@ZippyandOllie]

wait, so why are the 4.1 neutral again?

[@shanikhan00]

Because they were COO(-) to begin with. 4.1 makes it larger than the pH of 3.5, therefore they will gain an H and become COOH (neutral)

[@nurfatiniafiqah1952]

Good ... its helpful .. so we must memorize all the structures of amino acids? OMG ..

[@123Rukhsaar]

isn't it 2 similar points take an average of them in example 2 wouldn't it be (8.0+8.3)/2=8.15?

[@soniproductionz]

+Rukhsaar A Close, it is similar points around the average pH at which the net charge is 0. In example 2 that was pH of 7 so similar points around that pH of 7 would be 8.0 and 4.1.

[@rahimshah5859]

very helpfull

[@Dezzy7model]

I didn't understand in the second example the S_H part why \ how is it neutral when others are gaining or losing? besides it's pka value is 8.3 which is above 7 the pH value.!!!.??? pls help a broda out... 😱😨😤😡😫😓😭

[@midgetking101]

+Dezzy7model Since you are asking, I assume you haven't taken organic chemistry yet. There is a way to determine just by how many bonds it has. For this video, for sulfur specifically, it's probably just easier to remember that R-S-H will not be charged while R-S will be negative.

[@Dezzy7model]

+Yegor Rowan if that's the case then it should be -1 not neutral or 0??? but thanks for the reply. actually I took organic chemistry but we didn't cover sulphur affiliated stuffs.

[@thepowerofpositivity8075]

+Dezzy7model Hi! Jus follow the simple rule if (1) the compound is a weak acid then the protonated form will be uncharged (AH) and the deprotonated form (A) will be negatively charged;(2) the compound is a weak base then the protonated form (BH) will be positively charged and the deprotonated form (B) will be uncharged. The tendency a compound will donate or accept proton depends on pH of the solution. Hope this help!

[@Dezzy7model]

+Mien Le Am grateful for the response... but Can you connect it to my sulphur questions? because am guessing cysteine is a weak acid...

[@thepowerofpositivity8075]

Hi there again. Nothing new my friend the same rule as I shared with you. First of all, let's review the Henderson-Hasselbalch equation because this will help you clear the concept: pH = pKa + log [deprotonated "absence of "H"] / [protonated "presence of H"] so if a compound is (1) a weak acid: pH = pKa + log [A neg charge] / [AH no charge] or (2) a weak base: pH = pKa + log [B no charge] / [BH pos charge] Now apply this equation for Cysteine in the video: pH: 7 pKa: 8.3 (to simplify the math say it is 8) plug these values into the equation (1) for weak acid because we agree Cysteine is a weak acid 7 = 8 + log [A neg charge]/[AH no charge] minus 8 for both sides and rearrange: log [A neg charge]/[AH no charge] = -1 get rid of log both sides [A neg charge] / [AH no charge] = 10 to the power of negative 1 Rewrite: [A neg charge] / [AH no charge] = 1/10 or: [A neg charge] : [AH no charge] = 1 : 10 so obviously you can see that the concentration of AH (no charge or neutral form) is greater than A (neg charge form). This means at pH 7, Cysteine has tendency to be neutral or no charge.

[@superoxidedismutase5757]

Explanation was so good I raped the replay button.