Anthraniloyl-tyrosine 411 as a spectroscopic probe of fatty acid binding to human serum albumin.

N Hagag,R A Mcpherson,E R Birnbaum,D W Darnall

doi:10.1016/s0021-9258(18)91026-4

Abstract

Reaction of human serum albumin with p-nitrophenylanthranilate results in transesterification of the anthraniloyl group to tyrosine 411. Titration of anthraniloyl-Tyr-411-albumin with long chain or short chain fatty acids produces marked changes in the absorption and fluorescence spectra of the anthraniloyl moiety as fatty acids bind in the channel near it. It appears that the anthraniloyl group is a very sensitive probe that can follow binding of small molecules at the 3-AB subdomain of human serum albumin.

Highlights

Binding to Human Serum governing the six sites, serve as a probefor the relative binding affinity of this site compared to the othe5rsites
Titration of anthran- pronounced changes in both the absorption andfluorescence iloyl-Tyr-411-albumin with longchain or short chain spectra of the anthranilated protein due to faattcyid binding. fatty acidsproduces marked changes in the absorption we describe the experiments which indicate that anand fluorescence spectra of the anthraniloyl moiety as thraniloyl-HSA is anextremely sensitive probe for following fatty acids bind in the channel near it
It appears that fatty acid or other small molecule binding to the subdomain the anthraniloyl group is a very sensitive probe that 3-AB site (as defined by Brown and Shockley [3]) of human can follow binding of small molecules at the 3-AB serum albumin

Summary

MATERIALS ANDMETHODS

It iswell established thatHSA’ is themajor carrier of fatty dissolved in methanol and desired aliquowtsere introducedinto small acids in the plasma. When excited which could act as effective binding pockets for fatty acids. Koh and Means [4] have shown that asingle mole of a tered at 350 nm, is composed of two transitions, one short chain (C8 or Clo) fatty acid inhibited the acylation of of which exhibits a negative Cotton effect centered near 340. The effect of palmitic acid is to that Tyr-411 is either located close to one of the 6 channels increase the intensity of the 390 nm absorption band at the proposed as a fatty acid bindingsite, or thatTyr-411 is expense of the 340 nm absorption band. According to the Brown model of HSA, the first 3 mol of a long chain ’fatty acid should bind most strongly a t sites 1-C, 2-C, and 3-C,whereasupon further addition of fatty acid,.

Spectroscopic Probeof Fatty Acid Binding to Serum Albumin

Findings

The longest chainlengthfatty acidsproduce the largest