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alpha-Lactalbumin, Engineered to be Non-native and Inactive, Kills Tumor Cells when in Complex with Oleic Acid: A new biological function resulting from partial unfolding.

Pettersson, Jenny LU ; Mossberg, Anki LU ; Trulsson, Maria LU ; Yong, Yeon Joong LU ; Min, Soyoung; Lim, Yoongho; O'Brien, John E; Svanborg, Catharina LU and Mok, Ken LU (2009) In Journal of Molecular Biology 394(5). p.994-1010
Abstract
HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a tumoricidal complex consisting of partially unfolded protein and fatty acid, and was first identified in casein fractions of human breast milk. The complex can be produced from its pure components through a modified chromatographic procedure where pre-applied oleic acid binds with partially-unfolded alpha-lactalbumin on the stationary phase in situ. Because native alpha-lactalbumin itself cannot trigger cell death, HAMLET's remarkable tumor-selective cytotoxicity has been strongly correlated with the conformational change of the protein upon forming the complex, but whether a recovery to the native state subsequently occurs upon entering the tumor cell is yet unclear. To this... (More)
HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a tumoricidal complex consisting of partially unfolded protein and fatty acid, and was first identified in casein fractions of human breast milk. The complex can be produced from its pure components through a modified chromatographic procedure where pre-applied oleic acid binds with partially-unfolded alpha-lactalbumin on the stationary phase in situ. Because native alpha-lactalbumin itself cannot trigger cell death, HAMLET's remarkable tumor-selective cytotoxicity has been strongly correlated with the conformational change of the protein upon forming the complex, but whether a recovery to the native state subsequently occurs upon entering the tumor cell is yet unclear. To this end, we utilize a recombinant variant of human alpha-lactalbumin in which all eight cysteine residues are substituted for alanines (rHLA(all-Ala)), rendering the protein non-native and biologically inactive under all conditions. The HAMLET analogue formed from the complex of rHLA(all-Ala) and oleic acid (rHLA(all-Ala)-OA) exhibited equivalent strong tumoricidal activity against lymphoma and carcinoma cell lines, and was shown to accumulate within the nuclei of tumor cells, thus reproducing the cellular trafficking pattern of HAMLET. In contrast, the fatty acid-free rHLA(all-Ala) protein associated with the tumor cell surface but was not internalized and lacked any cytotoxic activity. Structurally, whereas HAMLET exhibited some residual native character in terms of NMR chemical shift dispersion, rHLA(all-Ala)-OA showed significant differences to HAMLET, and in fact was found to be devoid of any tertiary packing. The results identify alpha-lactalbumin as a protein with strikingly different functions in the native and partially unfolded states. We posit that partial unfolding offers another significant route of functional diversification for proteins within the cell. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Molecular Biology
volume
394
issue
5
pages
994 - 1010
publisher
Elsevier
external identifiers
  • wos:000272952400017
  • pmid:19766653
  • scopus:70450224072
ISSN
1089-8638
DOI
10.1016/j.jmb.2009.09.026
language
English
LU publication?
yes
id
2757875c-d2bf-42a6-ac12-3375b425d651 (old id 1483319)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/19766653?dopt=Abstract
date added to LUP
2009-10-07 12:35:11
date last changed
2017-08-13 03:40:52
@article{2757875c-d2bf-42a6-ac12-3375b425d651,
  abstract     = {HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a tumoricidal complex consisting of partially unfolded protein and fatty acid, and was first identified in casein fractions of human breast milk. The complex can be produced from its pure components through a modified chromatographic procedure where pre-applied oleic acid binds with partially-unfolded alpha-lactalbumin on the stationary phase in situ. Because native alpha-lactalbumin itself cannot trigger cell death, HAMLET's remarkable tumor-selective cytotoxicity has been strongly correlated with the conformational change of the protein upon forming the complex, but whether a recovery to the native state subsequently occurs upon entering the tumor cell is yet unclear. To this end, we utilize a recombinant variant of human alpha-lactalbumin in which all eight cysteine residues are substituted for alanines (rHLA(all-Ala)), rendering the protein non-native and biologically inactive under all conditions. The HAMLET analogue formed from the complex of rHLA(all-Ala) and oleic acid (rHLA(all-Ala)-OA) exhibited equivalent strong tumoricidal activity against lymphoma and carcinoma cell lines, and was shown to accumulate within the nuclei of tumor cells, thus reproducing the cellular trafficking pattern of HAMLET. In contrast, the fatty acid-free rHLA(all-Ala) protein associated with the tumor cell surface but was not internalized and lacked any cytotoxic activity. Structurally, whereas HAMLET exhibited some residual native character in terms of NMR chemical shift dispersion, rHLA(all-Ala)-OA showed significant differences to HAMLET, and in fact was found to be devoid of any tertiary packing. The results identify alpha-lactalbumin as a protein with strikingly different functions in the native and partially unfolded states. We posit that partial unfolding offers another significant route of functional diversification for proteins within the cell.},
  author       = {Pettersson, Jenny and Mossberg, Anki and Trulsson, Maria and Yong, Yeon Joong and Min, Soyoung and Lim, Yoongho and O'Brien, John E and Svanborg, Catharina and Mok, Ken},
  issn         = {1089-8638},
  language     = {eng},
  number       = {5},
  pages        = {994--1010},
  publisher    = {Elsevier},
  series       = {Journal of Molecular Biology},
  title        = {alpha-Lactalbumin, Engineered to be Non-native and Inactive, Kills Tumor Cells when in Complex with Oleic Acid: A new biological function resulting from partial unfolding.},
  url          = {http://dx.doi.org/10.1016/j.jmb.2009.09.026},
  volume       = {394},
  year         = {2009},
}