Dear All,
As we all know that lower Kd (micro-molar/nano-molar range) are considered as good interaction. But how one can appreciate the relative concept of this strength when I am measuring a single pair of protein-protein interaction. Please provide your valuable remarks on this. Thank you in advance.
Prem
Amended on 5 May 2019
This seems to be an interesting question, since multimer proteins such as IgM and multimer human serum biotinidase increase the affinity to substrate (please see file; The Fascio effect).
I have found that Km and Ki are important parameters in amidase and protease reactions (please see files; J Chrom B Rat BIN LIP Km and Enk serumBIN). Thus, we have firstly found that Ki value (Kip value) of liver biotinidase is increased in the cancer tissue (please see file; Anticancer Res).
I am now considering that one protein or subunit protein is considered as enzyme and another protein is considered as substrate/product subunit, and the Km and Ki values can be obtainable.
Since glycochains of enzyme have unexpectedly important role in the Km and Ki values, analysis of glycoprotein-glycoprotein interaction becomes to be an interesting issue (please see files; Wide rage of Biotin and JMBT Alopecia). Then, HPLC method may become to be the powerful tool in the research of glycoprotein-glycoprotein interactions.
By the way, Avidin is lipoic acid binding protein, since Avidin binds stronger to lipoate than to biotin as assessed by affinity chromatography (HPLC-trypsin-treated Avidin affinity chromatography) (please see file; Lipoic acid Avidin).
Further, nature has no Normal distribution or Gaussian distribution. Then, non-parametric statistical analysis is indispensable in Biochemistry.
Furthermore, since protein and glycoprotein change the affinity to substrate or ligand via the total chemical structure, old chemical idea of Kd should not be utilized in biochemistry.
"Strong" and "weak" are relative, and hence subjective, terms. A bond would certainly be called weak if the bond energy is in the order of the kinetic energy of molecules under Brownian motion, about 2 kT, where k is the Boltzmann constant and T the absolute temperature. On the other extreme, the strongest known non-covalent bond is that between avidin and biotin, KD ~ e-15 M. Calculated per molecule, Delta G = kT ln(KD) ~ 34 kT (recall that k = R/Na).
The strength of a given interaction can be judged through the association constant K or the dissociation constant Kd. Very roughly, and taking 1 M as the reference standard state concentration:
- Low affinity: Kd larger than 10-4 (> 100 microM)
- Moderate affinity: Kd between 10-4 and 10-7 (100 microM - 100 nM)
- Low affinity: Kd smaller than 10-7 (< 100 microM)
However, people working on in different fields may have different considerations. Thus, a Kd of 10-6 (1 microM) can be considered as high affinity in metabolism regulation, while it can be considered a low affinity in antibody design.
And this is related to another way to judge the strength of an interaction which takes into account the potential concentrations of the interacting molecules. Then, a Kd of 10-6 (1 microM) has different physiological consequences if the interacting molecules are present at mM (complex predominates), microM (complex and free molecules at comparable molar fractions) or nM (free molecules predominate) concentrations.
Thank you so much to all for your valuable time and suggestions. I really appreciate it. Thank you once again.
A strong interaction is indeed in the nM range. I seem to remember any potential drug was borderline ineffective in the high uM. The dose would be in grams.
That being said, I was tasked to get the enzyme I was working on to the lowest Kd manipulating; pH, salt, cofactors and co-substrate concentrations. Were these physiologically relevant (?) see Adrian Velazquez-Campoy
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